`_#P#` @@@ @@@@p6s0VDr#@#p EN DB #Q+ /  7q  aF   3 zEHH08#3 (PUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUUU<HH0 dHt8lLsLs _s c,fEgCEiBCwBCyCEzEECEBCBCCEEECEBCBCCEE,ECEBCBCCEEECEBCBCCEEPIKIIKI ( Dri|*|DtCBAbu-Zeid19967O Adams1998V Adams1998I Afare1993[ Aikawa19855C Aikawa19966S Aikawa19988\ Aikawa19989B Akida1996 Akoglu1987 Anand1987 Anders19878 Anders19889 Anders19909! Andrysiak1984N Arnot1999 Atkins19897@ Bayoumi1997 Bianco19877]Blackman1998V Blair1998C Blanton1996GBonnefoy19949HBonnefoy1994jF Borre1995 Brown1987 Brown1988 Brown1991J Brown1991Y Bushell1986 Bushell1988 Bushell1988BBygbjerg19967!Campbell1984&Campbell1984P Capiod19999G Carcy1994 Case19878NCavanagh1999L Certa1990 Chan19877 Chang1987 Cheng1992A Cheng1997 Chizzolini1984 Chizzolini1985n Clark1987_ Collins1999Y Cooper19866 Cooper19889 Cooper19889 Cooper19897 Cooper19909 Cooper19921E Cooper19959A Cooper19979 Coppel19877 Coppel19889 Coppel1991Crewther1987Crewther1990Culvenor1987Culvenor1990I Dalsgaard1993RDaniel-Ribeiro19989; Decrind1986+ Dedet1984S Del Rio1998^ Deloron1999Delplace19888L Delves19909Donachie1992rH Doury1994\ Doury1998^ Dubois19999 Dubremetz1988H Dubremetz1994; Dyal1986NElhassan1999KEtlinger19903 Etzion19919+ Fandeur1984G Fandeur1994 Fardoulys1988 Fardoulys1988RFerreira-da-Cruz19981^ Fievet19999>Fonjungo1998NFonjungo1999 Fortier1988! Franco19848WFranklin1993V Fraser19988% Freeman1981X Freeman1984[ Freeman1985C Fujioka1996S Fujioka1998\ Fujioka1998_ Fujioka1999\ Fusa1998 Gabra1985L Goman1990@ Greenwood1997GGrellier19944G Guillotte1994Harnyuttanakorn1992_ Hasnain1999Hauquitz19929IHeegaard1993BHeegaard19967Heidrich1992r\ Hienne1998_ Hoffman1999% Holder1981X Holder1984[ Holder1985 Holder19905J Holder19913F Holder19955Q Holder19989M Holder1999_Holloway1999& Howard1984 Howard19905 Howard19901  Howard19925W Howard1993D Howard1996? Howard1998R Howard1998i] Howard1998! Hudson19848 Hui1987I Hviid1993@ Hviid1997N Hviid1999 Ingram19888 Ingram19889 Irving19929?Jacobson1998RJacobson1998IJakobsen1993BJakobsen19967@Jakobsen1997U Jarra1998P Jarra1999W Jensen1993@ Jensen1997s Kan1987V Kappe1998  Keen19909J Keen19919F Keen19955M Keen19999 Kemp19878E Kennedy1995Y Kidson19866C King19969 Kramer19878_ Kumar1999@ Kurtzhals1997L Lahm19909 Lahm19919_ Lal1999* Lambert1981; Lambert1986GLe Scanf19944B Lemnge19969 Lewis1990 Limpaiboon19921 Loche1984+ Loche1984< Loche1984 Lockyer1990  Lustigman1988^ Luty1999i< Mach19848L Matile19909^ Mayombo1999^ Mbessi19999 McBride1987 McBride1992> McBride1998N McBride1999<McGarvey1984 GMercereau-Puijalon1994HMercereau-Puijalon1994 Merkli1984 Merkli1985B Mhina1996*Miescher1981^ Migot-Nabias1999! Miller19848^ Millet19999^ Minh19999@ Morris-Jones1997BMsangeni19967 Murray19919] Narum19989Ndengele19939I Nkrumah1993O Noe1998V Noe1998M Ogun19999ROliviera de Ferreira1998F Owen19955M Owen1999 Perkins1988: Perkins1990 Perkins1991 Perkins1991  Perkins1992* Perrin1981 Perrin1984+ Perrin1984e< Perrin19848 Perrin1985DPeterson19969 Playfair1990\Pradines1998U Preiser1998P Preiser1999EPrescott19959APrescott19977* Ramirez1981& Reese1984 Reese1990^ Renaut19999\ Ricard1998 Richle19841 Richle19851? Rickel19989R Rickel19988K Ridley19903L Ridley19909 Ridley199191 Ridley1991 Ridley19929I Ridley19939B Ridley19969@ Ridley19977@ Riley1997^Ringwald19991 Roger1988H Roger1994B Ronn19966N Roper1999+ Roussilhon1984B Ruta19966_ Sacci1999B Salum1996 Sam-Yellowe1988: Sam-Yellowe1990 Sam-Yellowe19917 Sam-Yellowe19928 Sam-Yellowe19939 Sam-Yellowe1993C Sam-Yellowe1996S Sam-Yellowe1998Y Saul19868 Saul1988 Saul19898 Saul19929E Saul19959T Saul19959A Saul19979K Scaife19909L Scaife19909 Scaife19919R Schmidt1998Y Schofield1986 Schofield1988GSchrevel19944ZSchulman19856<Sheybani1984 _ Shi1999 Shio1988w Siddiqui19878 Silva1990J Sinha1991F Sinha1995M Sinha1999 Smart1984P Snounou1999T Spring19955& Stanley1984 Stenzel1988Z Stewart1985 Stocker1985 Stowers1992E Stowers1995T Stowers1995A Stowers1997I Stuber19939> Stuber19989E Stueber1995K Takacs19903L Takacs19909E Takacs19959 Takcs19919 Tam1987A Taylor19977Q Taylor19989ITheander1993@Theander19977NTheander1999? Thurman1998R Thurman1998] Thurman1998Tronchin19888[ Uni1985Y Upcroft1986^Vaillant19991Z Vanderberg1985 Vernes19888Q Werner19989_ Wohlhueter1999BLWoollett19903S Yakubu19988 Yamaga19878C Yang19961S Yang19989im, G. C. Dourado, H. V. Wirth, D.F. 1986LESpecific DNA probe for the diagnosis of Plasmodium falciparum malariaScience 231 1434-1436^ 0780 Ref11h:3Barker, Rh Jr Suebsaeng, L. Rooney, W. Wirth, D. F.o}Detection of Plasmodium falciparum infection in human patients: a comparison of the DNA probe method to microscopic diagnosisAm J Trop Med Hyg 1989413C 266-72piBarker, Rh Jr Brandling, Bennett Ad Koech, D. K. Mugambi, M. Khan, B. David, R. David, J. R. Wirth, D. F.DD>Plasmodium falciparum: DNA probe diagnosis of malaria in Kenya Exp Parasitol  1989693 226-33&Barnwell, J. W. Desowitz, R. S. |vStudies on parasitic crisis in malaria: I. Signs of impending crisis in Plasmodium berghei infections of the white ratAnn Trop Med Parasitol 197771 429429-43302+Barnwell, J. W. Howard, R. J. Miller, L. H. `YInfluence of the spleen on the expression of surface antigens on parasitized erythrocytesCiba Found Sympn 198394 117]117-136:3Barnwell, J.W. Howard, R.J. Coon, H.G. Miller, L.H.$ 1983Splenic requirement for antigenic variation and expression of the variant antigen on the erythrocyte membrane in cloned Plasmodium knowlesi malariaInfect Immunity40985-994 0183 Ref116/Barnwell, J. W. Ockenhouse, C. F. Knowles, D.M.^ 1985Monoclonal antibody OKM5 inhibits the in vitro binding of Plasmodium falciparum-infected erythrocytes to monocytes, endothelial, and C32 melanoma cells  J Immunol    f11F@Barnwell, J. W. Asch, A. S. Nachman, R. L. Yamaya, M. Aikawa, M.A human 88-kD membrane glycoprotein (CD36) functions in vitro as a receptor for a cytoadherence ligand on Plasmodium falciparum-infected erythrocyteso J Clin Invest 198984 7659765-772(!Barnwell, J. W. Wertheimer, S. P..\UPlasmodium vivax: merozoite antigens, the Duffy blood group, and erythrocyte invasionnProg Clin Biol Res 1989 313n1           BR. AP(Theander, T.p(Adams, J. H.(McBride, J. @(8!HN }H8`cK>`888H;hy``dH;mM`888H;hQ``dhH;m%`H8}8!@8`N !8`~}~PHK`9`,A9`a},8@8~97178269Jakobsen, P. H. Lemnge, M. M. Abu-Zeid, Y. A. Msangeni, H. A. Salum, F. M. Mhina, J. I. Akida, J. A. Ruta, A. S. Ronn, A. M. Heegaard, P. M. Ridley, R. G. Bygbjerg, I. C.Immunoglobulin G reactivities to rhoptry-associated protein-1 associated with decreased levels of Plasmodium falciparum parasitemia in Tanzanian children Age Factors Analysis of Variance Animal Antibodies, Protozoan/biosynthesis Antigens, Protozoan/immunology Child Child, Preschool Enzyme-Linked Immunosorbent Assay Human IgG/*biosynthesis IgM/biosynthesis Infant Malaria, Falciparum/epidemiology/*immunology/parasitology Parasitemia/epidemiology/*immunology/parasitology Plasmodium falciparum/*immunology Prevalence Protozoan Proteins/*immunology Recombinant Proteins/immunology Support, Non-U.S. Gov't Tanzania/epidemiologyIn the Muheza region of Tanzania, an area with holoendemic malaria, the proportion of responders with IgG enzyme-linked immunosorbent assay reactivities to recombinant rhoptry-associated protein-1 (rRAP-1) as well as IgG reactivities to a repeat region of the acidic-basic repeat antigen (ABRA) increased with age. The proportion of responders with IgM reactivities to rRAP-1 increased with age in the first three decades. However, levels of IgG reactivities to rRAP-1 did not increase with age, indicating high levels of reactivities among young children. High P. falciparum densities were only detectable in children less than five years of age; in this group the proportion of IgG responders to rRAP-1 and to the ABRA repeat region was low but levels of IgG reactivities to rRAP-1 were inversely correlated with parasite density, suggesting that immune recognition of the antigen may be associated with resistance to infection. On the other hand, levels of IgG reactivities to the repeat region of ABRA increased with parasite densities in children 1-4 years of age. Two different profiles of IgG reactivities to rRAP-1 and to ABRA are detectable in young Tanzanian children and the Ig reactivities against rRAP-1 may be a component of the immune reactions restricting parasite multiplication.Am J Trop Med Hyg 1996556 642-6PUr+* |Perkins, M. E. 19922,Rhoptry organelles of apicomplexan parasitesParasitol Today8 28-32>8Perrin, L. H. Ramirez, E. Lambert, P. H. Miescher, P. A.XQInhibition of P. falciparum growth in human erythrocytes by monoclonal antibodies. Nature 1981 289^301-303n Ref11HBPerrin, L. H. Merkli, B. Loche, M. Chizzolini, C. Smart, J. Richle 1984lfAntimalarial immunity in Saimiri monkeys. Immunization with surface components of asexual blood stages J Exp Med 160441-451 0108 Ref11 F?Perrin, L. H. Loche, M. Dedet, J. P. Roussilhon, C. Fandeur, T.s\VImmunization against Plasmodium falciparum asexual blood stages using soluble antigensClin Exp Immunol 198456 67-72 Ref11TNPerrin, L. H. Merkli, B. Gabra, M. S. Stocker, J. W. Chizzolini, C. Richle, R.piImmunization with a Plasmodium falciparum merozoite surface antigen induces a partial immunity in monkeysc J Clin Invest9 198575 1718-1721 Ref11 zthttp://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://www.idealibrary.com/cgi-bin/links/citation/0014-4894/89/5098264651Preiser, P. R. Jarra, W.Plasmodium yoelii: differences in the transcription of the 235-kDa rhoptry protein multigene family in lethal and nonlethal linesi Amino Acid Sequence Animal Antibodies, Monoclonal/immunology/therapeutic use Antigens, Protozoan/chemistry/genetics/immunology DNA, Protozoan/chemistry Female Gene Expression Immunization, Passive Malaria/immunology/parasitology/therapy Mice Mice, Inbred BALB C Molecular Sequence Data *Multigene Family Plasmodium yoelii/*genetics/pathogenicity Polymerase Chain Reaction Protozoan Proteins/chemistry/*genetics/immunology RNA, Protozoan/genetics Sequence Alignment Specific Pathogen-Free Organisms *Transcription, GeneticiWe have compared the transcription of the 235-kDa rhoptry protein (p235) multigene family in the lethal (YM) and nonlethal (17X) lines of the rodent malaria parasite Plasmodium yoelii. This protein is thought to be involved in erythrocyte invasion by the parasite. Using a PCR- based approach we demonstrated that both lines have similar p235 families. However, RT-PCR analysis revealed that this similarity is not evident at the level of transcription, with the lethal line not transcribing a whole subset of its p235 gene repetoire. Specific anti- p235 immune pressure induces differences in invasion properties of the lethal line; we were, however, unable to detect any changes in the transcription pattern of the p235 genes associated with this event. Exp Parasitol 1998891 50-7992314966/Preiser, P. R. Jarra, W. Capiod, T. Snounou, G.dlfA rhoptry-protein-associated mechanism of clonal phenotypic variation in rodent malaria [see comments]Animal Clone Cells Erythrocytes/parasitology Female Genes, Protozoan Malaria/*parasitology Mice Mice, Inbred BALB C Multigene Family Phenotype Plasmodium yoelii/*genetics/physiology Polymerase Chain Reaction Protozoan Proteins/*genetics/physiology Reverse Transcriptase Polymerase Chain Reaction Rodent Diseases/parasitology RNA, Protozoan/genetics Transcription, Genetic *Variation (Genetics) zsThe recognition and invasion of host cells are mediated by components of the apical complex of the ookinete, sporozoite and merozoite stages of Plasmodium parasites. The paired rhoptries (organelles involved in host-cell recognition) in the apical complex contain many proteins of as-yet unknown function. In the rodent malaria agent P. yoelii yoelii, a multigene family codes for merozoite rhoptry proteins of relative molecular mass 235,000 (p235 proteins); these proteins are thought to determine the subset of erythrocytes that the parasites invade. Further support for this idea came from the identification of a region in p235 with weak but significant homology to reticulocyte-binding protein-2 of P. vivax and the demonstration that at least one p235 member binds to the erythrocyte surface membrane. Here, using single, micromanipulated P.y.yoelii parasites, we describe a new mechanism of gene expression by which the merozoites originating from a single schizont each express a distinct member of this multigene family. We propose that this new type of clonal phenotypic variation provides the parasite with a survival strategy in the mammalian host; this strategy contributes to the observed chronicity of malarial infections. This phenomenon is genetically and functionally distinct from classical antigenic variation, which is mediated by the var multigene family of P. falciparum.@ Nature 1999 398) 6728 618-22  Authors JournalsKeywords   Kowe,mw(. Fujioka, HKpkawa, M. Yang, J. C. Yakubu, Z.w(Yakubu, Z.. wx0(8!@N }wx0(`?$w8~;8HM[!`8~HM^`H(}8h8~HM]`8}N 8~HMZ` b,,~ 0($49@ ^8H(}8h8 `K78}N <~<HLD@P^TH(}8h8<`K7u8}N 8~X8 HL)`0ctHUc8HH}8h8~XHL)`8}N }8h8<`H+8}N }8h8 `H+8}N }8h8~HM\`8}N }891088206:3Ridley, R. G. Takacs, B. Etlinger, H. Scaife, J. G.HRKA rhoptry antigen of Plasmodium falciparum is protective in Saimiri monkeys vpAnimal Antigens, Protozoan/*immunology/isolation & purification Blotting, Western Chromatography, Affinity Electrophoresis, Polyacrylamide Gel Fluorescent Antibody Technique Immune Sera/immunology Malaria/*prevention & control Plasmodium falciparum/*immunology Protozoan Proteins/*immunology/isolation & purification *Protozoan Vaccines Saimiri Support, Non-U.S. Gov'tD>A non-polymorphic antigen associated with the rhoptry organelles of Plasmodium falciparum has been purified by immunoaffinity chromatography. The antigen, RAP-1 (rhoptry associated protein-1), which is defined by monoclonal antibodies which inhibit parasite growth in vitro, is a multi-component antigen consisting of four major proteins of 80, 65, 42 and 40 kDa and two minor proteins of 77 and 70 kDa. These proteins were electro-eluted from preparative sodium dodecyl sulphate polyacrylamide gels and protected Saimiri sciureus monkeys from a lethal blood-stage infection of P. falciparum malaria. Sera from the protected animals recognized only proteins of the RAP-1 antigen when used to probe a Western blot of total parasite protein extract, confirming that RAP-1 is responsible for eliciting the protective immune response. Parasitology 1990101 Pt 2 187-92 :_YrS 9(!Sam-Yellowe, T.Y. Ndengele, M. M. 1993TMMonoclonal antibody epitope mapping of Plasmodium falciparum rhoptry proteins Exp Parasitol761  46-58|uhttp://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://www.idealibrary.com/cgi-bin/links/citation/0014-4894/89/27198339327VOSam-Yellowe, T. Y. Del Rio, R. A. Fujioka, H. Aikawa, M. Yang, J. C. Yakubu, Z.Isolation of merozoite rhoptries, identification of novel rhoptry- associated proteins from Plasmodium yoelii, P. chabaudi, P. berghei, and conserved interspecies reactivity of organelles and proteins with P. falciparum rhoptry-specific antibodiesAnimal Antibodies, Monoclonal/immunology Antibodies, Protozoan/immunology Antigens, Protozoan/*analysis/chemistry/immunology Blotting, Western Cell Fractionation Centrifugation, Density Gradient Electrophoresis, Polyacrylamide Gel Epitopes/analysis/chemistry/immunology Female Fluorescent Antibody Technique, Indirect Immune Sera/immunology Male Mice Microscopy, Immunoelectron Molecular Weight Organelles/chemistry/immunology/ultrastructure Plasmodium berghei/*chemistry/immunology/ultrastructure Plasmodium chabaudi/*chemistry/immunology/ultrastructure Plasmodium yoelii/*chemistry/immunology/ultrastructure Precipitin Tests Protozoan Proteins/*analysis/chemistry/immunology Silver Staining Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.Rhoptries were isolated from merozoites of P. yoelii (17 XL), P. chabaudi adami and P. berghei (K-173), using sucrose gradient density centrifugation. Mouse antisera was prepared against the organelles and characterized. Antibodies specific for a known P. yoelii rhoptry protein were used to identify gradient fractions containing rhoptries and electron microscopy was used to confirm rhoptry enrichment and organelle morphology. Western blotting analysis of the gradients with organelle-specific antisera from each species, revealed several major cross-reactive interspecies protein bands of approximately 235, 210, 180, 160/170, 140, and 96-110 kDa, predominantly in densities of 1.12 and 1.15 g/ml. The parasite origin of the proteins was verified by immunoprecipitation, and reactive epitopes localized to the rhoptries by IEM. By Western blotting antisera specific for P. falciparum rhoptries reacted with protein bands of approximately 96-110 kDa in schizont extracts, and gradient fractions of density 1.12 and 1.15 g/ml from all three rodent malaria species, as well as with the rhoptries in P. yoelii, P. chabaudi, and P. berghei merozoites by IEM. We conclude that the three rodent malaria species and P. falciparum share conserved interspecies epitopes. Exp Parasitol 1998893 271-84VPSaul, A. Cooper, J. Hauquitz, D. Irving, D. Cheng, Q. Stowers, A. Limpaiboon, T. 1992LEThe 42-kilodalton rhoptry-associated protein of Plasmodium falciparumMol Biochem Parasitol50139-15086174890VPSchofield, L. Bushell, G. R. Cooper, J. A. Saul, A. J. Upcroft, J. A. Kidson, C.A rhoptry antigen of Plasmodium falciparum contains conserved and variable epitopes recognized by inhibitory monoclonal antibodiesAnimal Antibodies, Monoclonal/immunology Antigens, Protozoan/analysis/*immunology Brazil Electrophoresis, Polyacrylamide Gel Epitopes/analysis/*immunology Erythrocytes/immunology/*parasitology Fluorescent Antibody Technique Human Immunosorbent Techniques Malaria/immunology/parasitology Organoids/immunology Papua New Guinea Plasmodium falciparum/growth & development/*immunology/ultrastructure Radioimmunoassay Support, Non-U.S. Gov't ThailandyFour monoclonal antibodies produced against Plasmodium falciparum recognize an antigen in merozoites that is localized in rhoptries, as judged by a punctate, double dot fluorescence pattern. All four antibodies bound to the same affinity purified antigen in a two site immunoradiometric assay. Immunoprecipitation of antigen by monoclonal antibody followed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis yielded protein bands of 80, 66 and 42 kDa. Western blotting gave bands of 80 and 66 kDa only with three of the antibodies: the fourth did not blot. Based on protease inhibitor data the 66 kDa band is considered to be a cleavage product of the 80 kDa band, but the 42 kDa band does not appear to derive from the latter and may be a coprecipitation product. This group of antigens labels with both [35S]methionine and [3H]histidine. Two of the monoclonal antibodies inhibited merozoite invasion of erythrocytes. One of these inhibitors recognizes a variable epitope, whereas the second recognizes a highly conserved epitope present in all 106 primary isolates of P. falciparum tested from Brazil, Thailand and Papua New Guinea.Mol Biochem Parasitol  1986182 183-95}Shi, Y.P. Hasnain, S.E. Sacci, J.B. Holloway, B.P. Fujioka, H. Kumar, N. Wohlhueter, R. Hoffman, S.L. Collins, W.E. Lal, A.A. 1999sImmunogenicity and in vitro protective efficacy of a recombinant multistage Plasmodium falciparum candidate vaccine   L a  Proc. Nat. Acad. Sci. USA964j 1615-1620VOProceedings of the National Academy of Sciences of the United States of AmericavpSiddiqui, W. A. Tam, L. Q. Kramer, K. J. Hui, G. S. Case, S. E. Yamaga, K. M. Chang, S.P. Chan, E.B.T. Kan, S-C.vpMerozoite surface coat precursor protein completely protects Aotus monkeys against Plasmodium falciparum malariaProc Natl Acad Sci U S A 198784 3014-3018@ Ref11(<*  V @ 98007083Jakobsen, P. H. Kurtzhals, J. A. Riley, E. M. Hviid, L. Theander, T. G. Morris-Jones, S. Jensen, J. B. Bayoumi, R. A. Ridley, R. G. Greenwood, B. M.Antibody responses to Rhoptry-Associated Protein-1 (RAP-1) of Plasmodium falciparum parasites in humans from areas of different malaria endemicityAdolescence Adult Aged Animal Antibodies, Protozoan/*immunology Antigens, Protozoan/immunology Child Child, Preschool Cross-Sectional Studies *Endemic Diseases Female Gambia/epidemiology Human Infant Kenya/epidemiology Longitudinal Studies Malaria, Falciparum/blood/epidemiology/*immunology Male Middle Age Plasmodium falciparum/*immunology Protozoan Proteins/*immunology Recombinant Fusion Proteins/immunology Sudan/epidemiology Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.ePlasma IgM and IgG antibody reactivities against the recombinant Plasmodium falciparum protein, Rhoptry Associated Protein-1 (rRAP-1) were measured by ELISA in individuals from Sudan, Indonesia, Kenya and The Gambia living in areas of different malaria endemicity. IgG and IgM reactivities to rRAP-1 increased with malaria endemicity. IgG reactivities were associated with spleen rates in Indonesia with high malaria endemicity while IgM reactivities were associated with spleen rates in Kenya with low malaria endemicity. IgG and IgM reactivities to rRAP-1 increased during acute episodes of P. falciparum malaria in Sudanese adults and IgG reactivities remained high one month after treatment in all adults tested. Antibody reactivities to rRAP-1 in Gambian children in the dry season were higher in children with parasitaemia than in children without detectable parasitaemia. Antibody reactivities were not associated with protection against clinical episodes in the following rainy season but higher antibody reactivities were detectable at the end of the rainy season. There was no difference in antibody reactivity to rRAP-1 between Gambian children with mild or severe malaria.Parasite Immunol 1997199  387-93hahttp://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://www.pnas.org/cgi/content/full/95/3/123098115903HBKappe, S. H. I. Noe, A. R. Fraser, T. S. Blair, P. L. Adams, J. H.LFA family of chimeric erythrocyte binding proteins of malaria parasitesAmino Acid Sequence Animal Antigens, Protozoan/chemistry Antigens, Surface/chemistry Carrier Proteins/chemistry/genetics/*isolation & purification Cell Adhesion Molecules/chemistry/genetics/*isolation & purification Chimeric Proteins/chemistry/genetics/isolation & purification Consensus Sequence Duffy Blood-Group System/chemistry/genetics/*isolation & purification DNA, Protozoan/isolation & purification Membrane Proteins/chemistry Mice Mice, Inbred BALB C Molecular Sequence Data Plasmodium berghei/*chemistry/*genetics Plasmodium yoelii/*chemistry/*genetics Protozoan Proteins/chemistry Receptors, Cell Surface/chemistry/genetics/*isolation & purification RNA Splicing Sequence Alignment Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. Transcription, GeneticoProteins sequestered within organelles of the apical complex of malaria merozoites are involved in erythrocyte invasion, but few of these proteins and their interaction with the host erythrocyte have been characterized. In this report we describe MAEBL, a family of erythrocyte binding proteins identified in the rodent malaria parasites Plasmodium yoelii yoelii and Plasmodium berghei. MAEBL has a chimeric character, uniting domains from two distinct apical organelle protein families within one protein. MAEBL has a molecular structure homologous to the Duffy binding-like family of erythrocyte binding proteins located in the micronemes of merozoites. However, the amino cysteine- rich domain of MAEBL has no similarity to the consensus Duffy binding- like amino cysteine-rich ligand domain, but instead is similar to the 44-kDa ectodomain fragment of the apical membrane antigen 1 (AMA-1) rhoptry protein family. MAEBL has a tandem duplication of this AMA-1- like domain, and both of these cysteine-rich domains bound erythrocytes when expressed in vitro. Differential transcription and splicing of the maebl locus occurred in the YM clone of P. yoelii yoelii. The apical distribution of MAEBL suggested localization within the rhoptry organelles of the apical complex. We propose that MAEBL is a member of a highly conserved family of erythrocyte binding proteins of Plasmodium involved in host cell invasion. Proc Natl Acad Sci U S A 1998953  1230-5>7Keen, J. Holder, A., Playfair, J. Lockyer, M. Lewis, A. 1990rkIdentification of the gene for a Plasmodium yoelii rhoptry protein. Multiple copies in the parasite genomeMol Biochem Parasitol`42241-246 0985 Ref114<6Lustigman, S. Anders, R. F. Brown, G. V. Coppel, R. L. 1988piA component of an antigenic rhoptry complex of Plasmodium falciparum is modified after merozoite invasionMol Biochem Parasitol30217-224 0622 Ref11D=McGarvey, M. J. Sheybani, E. Loche, M. P. Perrin, L. Mach, B. 1984Identification and expression in Escherichia coli of merozoite stage-specific genes of the human malarial parasite Plasmodium falciparumProc Natl Acad Sci U S A81 3690-3694 0496 MonashC Q A 97313167HAStowers, A. Taylor, D. Prescott, N. Cheng, Q. Cooper, J. Saul, A.rkAssessment of the humoral immune response against Plasmodium falciparum rhoptry-associated proteins 1 and 2 Adolescence Adult Age Factors Animal Antibodies, Bacterial/*blood Antibody Specificity Child Child, Preschool Human Infant Infant, Newborn Parasitemia/immunology Plasmodium falciparum/*immunology Protozoan Proteins/*immunology Support, Non-U.S. Gov'tNaturally occurring antibody responses to Plasmodium falciparum rhoptry- associated proteins 1 and 2 (RAP-1 and RAP-2) were measured with recombinant and parasite-derived forms of the antigens. For comparative purposes, responses to multiple forms of three other malarial antigens were also examined. The sera of 100 Papua New Guineans were screened for antibodies. Eighty-six and 82% of individuals over 30 years of age had antibodies that recognized parasite-derived RAP-1 and RAP-2, respectively. Importantly, we found that recombinant and native antigens share linear epitopes seen by the human immune system; thus, the recombinant proteins may be adequate human immunogens. However, antibodies affinity purified on recombinant RAP-1 reacted with other antigens in addition to parasite-derived RAP-1. Thus, the antigenicity of RAP-1 may have been overestimated previously. The recognition of RAP- 1 and RAP-2 correlated with age and with the recognition of recombinant forms of the ring-infected erythrocyte surface antigen, merozoite surface protein 1, and merozoite surface antigen 2 (MSA2) antigens. Antibodies to these antigens appear to be generated in response to the total exposure to malaria of the host. Antibodies to conserved regions of MSA2 had stronger correlations with both age and the recognition of other antigens than did the full-length recombinant MSA2 molecule. In contrast to results with the other antigens, there was no significant difference in the ages of individuals with a certain antibody titer to the full-length recombinant or parasite-derived MSA2 molecule, but antibodies to these two antigens did correlate with parasitemia. For all antigens tested, antibody levels after two infections can approach the peak levels of antibodies obtained in immune individuals.J Infect Immun 19976562329-38n984187650)Werner, E. B. Taylor, W. R. Holder, A. A.,jcA Plasmodium chabaudi protein contains a repetitive region with a predicted spectrin-like structureAmino Acid Sequence Animal Cloning, Molecular Dimerization DNA, Complementary DNA, Protozoan/analysis Erythrocytes/parasitology Fluorescent Antibody Technique Genes, Protozoan Leucine Zippers Malaria/*parasitology Mice Mice, Inbred BALB C Molecular Sequence Data Plasmodium chabaudi/*chemistry/genetics/isolation & purification Protein Structure, Secondary Protozoan Proteins/*chemistry/genetics/isolation & purification Rats Rats, Inbred F344 Repetitive Sequences, Amino Acid Spectrin/chemistry/genetics Support, Non-U.S. Gov'tcDNA and genomic DNA clones covering the entire open reading frame (ORF) for a Plasmodium chabaudi 96V protein were isolated. From the first ATG codon the intronless gene codes for a 229-kDa protein. Antisera raised against recombinant polypeptides coded by two different regions of the gene reacted with a 240/225-kDa doublet on Western blots of parasite extracts. In immunofluorescence studies the same sera detected the antigen at the apical end of the merozoite, possibly in rhoptry organelles. In Western blotting experiments the recombinant polypeptides were recognised by antibodies induced by natural infection. A 364-amino acid residue repetitive region, based on 32 11- mer repeats divided by two 6-mer repeats into three blocks, is located in the centre of the protein. Analysis of this repetitive region led us to propose a model in which each of the three units forms an alpha- helical coiled-coil triple-helix containing a possible leucine- histidine zipper. Each unit resembles in structure the units present in spectrin. The repeat region is flanked by predicted heptad based alpha- helical coiled-coil regions, and we propose that the protein forms a dimer. The 229-kDa protein has the overall character of a cytoskeletal protein. We have named the 229-kDa protein repetitive organellar protein (ROPE) and suggest that ROPE may be involved in the process of invasion, possibly by interacting with the erythrocyte cytoskeleton, and that the leucine histidine-zipper may be involved in molecular mimicry of spectrin.iMol Biochem Parasitol 1998942 185-9696355865VPYang, J. C. Blanton, R. E. King, C. L. Fujioka, H. Aikawa, M. Sam-Yellowe, T. Y.Seroprevalence and specificity of human responses to the Plasmodium falciparum rhoptry protein Rhop-3 determined by using a C-terminal recombinant proteinAnimal Antibodies, Protozoan/*immunology Antigens, Protozoan/*genetics/*immunology Cloning, Molecular Comparative Study Female Fetal Blood/immunology Human IgG/immunology Immunity, Maternally-Acquired Malaria, Falciparum/epidemiology/*immunology Plasmodium falciparum/*immunology Plasmodium vivax/immunology Pregnancy Protozoan Proteins/*genetics/*immunology Recombinant Proteins/immunology Serologic Tests Species Specificity Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.Rhoptry proteins participate in invasion of erythrocytes by malaria parasites. Antibodies to some of these proteins can inhibit invasion and partially protect monkeys from disease. To examine human serological responses to the 110-kDa component (Rhop-3) of the high- molecular-weight rhoptry protein complex, two cDNA clones corresponding to Rhop-3 were identified by immunologic screening. A recombinant protein representing the C-terminal one-third of the Rhop-3 was used to assess the seroprevalence to this protein in geographically isolated populations with different patterns of malaria transmission. The immunoglobulin G (IgG) positivity rate for the recombinant Rhop-3 in an enzyme-linked immunosorbent assay was 30% in an area of Papua New Guinea where malaria is holoendemic. In Kenya, the prevalence rates were 43 and 36%, respectively, in an area of hyperendemicity and an area of seasonal transmission. By contrast, rates of IgG seroprevalence to an extract of Gambian strain of Plasmodium falciparum were 48, 90, and 97% respectively, in these populations. In these areas, the pattern of antibody recognition of Rhop-3 is more similar (1.7-fold maximum difference) than the parasite extract (5-fold difference). The difference in seroresponses may represent antigenic polymorphism in different parasite strains, while their similarity for the Rhop-3 fragment may represent conservation of this protein. Recombinant- and parasite extract-specific IgG was not found in individuals infected only with Plasmodium vivax. Cross-reactivity was seen in the IgM assay. In Mombasa (Kenya), maternal and cord Rhop-3-specific IgG activities were similar. Fetal antigen-specific IgM reactivity was generally undetectable for all antigens.  Infect Immun 19966493584-91t ;H|6G!F95364832F?Borre, M. B. Owen, C. A. Keen, J. K. Sinha, K. A. Holder, A. A.XQMultiple genes code for high-molecular-mass rhoptry proteins of Plasmodium yoeliirzAnimal Base Sequence Chromosome Mapping Cloning, Molecular Comparative Study *Genes, Structural, Protozoan Mice Molecular Sequence Data Plasmodium yoelii/*genetics/immunology Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Protozoan Proteins/*genetics/immunology Sequence Alignment Sequence Homology, Nucleic Acid Support, Non-U.S. Gov't Variation (Genetics)tnWe have examined the number of genes coding for a group of high- molecular-mass rhoptry protein(s) in the malaria parasite Plasmodium yoelii, and studied variation in the gene family within the parasite's genome. A region of the genes was amplified using oligonucleotides based on conserved DNA sequences and the products cloned. The sequences could be divided into 7 groups by restriction-fragment-length polymorphism. Further variation was detected by sequence analysis; 11 different sequences were detected in the 16 clones analyzed. The genes in the family were distributed on 6 chromosomes probably at 9 or more loci.Mol Biochem Parasitol 199570 1-2 149-55 Brown, H. J. Coppel, R. L.BAn antigenic complex in the rhoptries of Plasmodium falciparumMol Biochem Parasitol28105-112 0633 Ref11 NGCampbell, G. H. Miller, L. H. Hudson, D. Franco, E. L. Andrysiak, P. M. 1984LFMonoclonal antibody characterization of Plasmodium falciparum antigensAm J Trop Med Hyg33 1051-1054 0128 Ref1195258079tnCarcy, B. Bonnefoy, S. Guillotte, M. Le Scanf, C. Grellier, P. Schrevel, J. Fandeur, T. Mercereau-Puijalon, O.d^A large multigene family expressed during the erythrocytic schizogony of Plasmodium falciparumAmino Acid Sequence Animal Antigens, Protozoan/immunology Babesia/genetics/immunology Base Sequence Cross Reactions DNA, Protozoan/analysis Erythrocytes/*parasitology Gene Expression *Genes, Protozoan Human Malaria/immunology Molecular Sequence Data Multigene Family/*genetics/immunology Plasmodium falciparum/*genetics/immunology/physiology Polymorphism (Genetics) Protozoan Proteins/genetics/immunology Rabbits Repetitive Sequences, Nucleic Acid Sequence Homology, Amino Acid Support, Non-U.S. Gov'tcWe report the identification of a large multigene family of Plasmodium falciparum using a clone isolated with a polyclonal antiserum raised to a Babesia divergens merozoite protein. The recombinant antigen reacted with human sera collected from individuals exposed to malaria. The deduced protein sequence contains a motif homologous to the consensus sequence of merozoite rhoptry proteins encoded by multigene families in several Babesia species. Antibodies raised to the recombinant protein reacted with a 60-kDa merozoite protein both on B. divergens and on P. falciparum immunoblots. The insert hybridized to a large number of fragments on P. falciparum Southern blots and to most chromosomes of the parasite. Specifically, approx. 3-kb RNAs were detected in 4-16- nucleus schizonts. Ten distinct cDNAs were isolated that differed in the size, position and number of restriction sites in the region homologous to the original genomic clone. With about 140 copies per haploid genome, this is the first large multigene family described in malaria parasites. The existence of a multigene family encoding proteins present in the invasive stage of malaria parasites suggests an important role in invasion and denotes a significant potential for generating diversity.eMol Biochem Parasitolc 1994682e 221-3360Clark, J. T. Anand, R. Akoglu, T. McBride, J. S. 1987zIdentification and characterisation of proteins associated with the rhoptry organelles of Plasmodium falciparum merozoites Parasitol Res73425-434 0790 Ref11 f`Cooper, J. A. Ingram, L. T. Bushell, G. R. Fardoulys, C. A. Stenzel, D. Schofield, L. Saul, A.J. 1988|vThe 140/130/105 kilodalton protein complex in the rhoptries of Plasmodium falciparum consists of discrete polypeptidesMol Biochem Parasitol29251-260 0748 Ref11(!Cooper, J. A. Atkins, A. Saul, A./f_N-terminal amino acid sequencing of the 105 kilodalton rhoptry antigen of Plasmodium falciparumMol Biochem Parasitolu 198933203-204]f`Coppel, R. L. Bianco, A. E. Culvenor, J. G. Crewther, P. E. Brown, G. V. Anders, R.F. Kemp, D.J. 1987HBA cDNA clone expressing a rhoptry protein of Plasmodium falciparumMol Biochem ParasitolT25 73-81 0414 Ref11HACrewther, P.E. Culvenor, J.G. Silva, A. Cooper, J.A. Anders, R.F. 1990nhPlasmodium falciparum: Two antigens of similar size are located in different compartments of the rhoptry Exp Parasitol@70193-206 0888 Ref1194294202RKDoury, J. C. Bonnefoy, S. Roger, N. Dubremetz, J. F. Mercereau-Puijalon, O.epjAnalysis of the high molecular weight rhoptry complex of Plasmodium falciparum using monoclonal antibodiesAnimal Antibodies, Monoclonal/*immunology Antigens, Protozoan/*analysis/chemistry/genetics/immunology Base Sequence Binding, Competitive DNA Primers/chemistry Enzyme-Linked Immunosorbent Assay Epitopes/analysis/immunology Fluorescent Antibody Technique Immunoblotting Mice Microscopy, Immunoelectron Molecular Sequence Data Molecular Weight Plasmodium falciparum/*chemistry/immunology Precipitin Tests Protozoan Proteins/*analysis/chemistry/genetics/immunology Recombinant Proteins/immunology Species Specificity Support, Non-U.S. Gov'tPJTwenty-one monoclonal antibodies, obtained after immunization of mice with erythrocytic stages of Plasmodium falciparum, produced a double dot image in IFA. Immunoelectronmicroscopy indicated that the mAbs reacted with the rhoptries. Rhoptries are pear-shaped apical organelles, believed to be involved in invasion of the host cell by the parasite. The mAbs all immunoprecipitated the high molecular weight antigen complex. Some mAbs recognized on immunoblots only 1 protein of this complex, whereas others reacted with RhopH1 and RhopH3, or RhopH2 and RhopH3 or with the 3 proteins. An additional antigen of 52 kDa was also recognized by some of the mAbs. The epitopes defined by the mAbs were present in most of the 40 P. falciparum strains or isolates studied by IFA. Interestingly, the mAbs also reacted with high titres on P. vivax and P. ovale, but produced images that did not indicate an apical location. The mAbs failed to react on the non-human malaria parasites studied, P. cynomolgi and P. inui. On P. berghei or P. chabaudi parasites, only 5 mAbs gave a positive reaction, labelling a large network outside the parasite. Finally, the mAbs did not react with P. falciparum sporozoites, indicating that the rhoptries of merozoites and sporozoites, the two invasive stages of the malaria life- cycle are equipped with distinct sets of proteins. Parasitology 1994 108 Pt 3 269-80("Dyal, R. Decrind, C. Lambert, P.H. 1986zComparison of asexual blood-stage antigens of Plasmodium falciparum recognized by antibody reagents from nine laboratories Bull W.H.O.64403-414IR ]|60Howard, R.F. Narum, D.L. Blackman, M. Thurman, J 1998Analysis of the processing of Plasmodium falciparum rhoptry-associated protein 1 and location of Pr86 to schizont rhoptries and p67 to free merozoites  3 Mol. Biochem. Parasitol.92111-122,&Molecular and Biochemical Parasitology98379791Jacobson, K. C. Thurman, J. Schmidt, C. M. Rickel, E. Oliviera de Ferreira, J. Ferreira-da-Cruz, M. F. Daniel-Ribeiro, C. T. Howard, R. F.A study of antibody and T cell recognition of rhoptry-associated protein-1 (RAP-1) and RAP-2 recombinant proteins and peptides of Plasmodium falciparum in migrants and residents of the state of Rondonia, BrazilJDAdolescence Adult Age Distribution Aged Animal Antibodies, Monoclonal/immunology Antibodies, Protozoan/*biosynthesis/immunology Antigens, Protozoan/chemistry/*immunology Brazil/epidemiology Child Cytokines/biosynthesis Female Human IgG/immunology IgM/immunology Lymphocyte Transformation Malaria, Falciparum/epidemiology/*immunology/parasitology Male Middle Age Plasmodium falciparum/chemistry/*immunology Protozoan Proteins/*immunology Recombinant Proteins/chemistry/immunology Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. T-Lymphocytes/*immunology Transients and MigrantsHumoral and cellular responses were examined among natives and migrants in an area of the Amazon region of Brazil. Rhoptry-associated protein-1 (RAP-1) and RAP-2 expressed in Escherichia coli expression systems, a peptide corresponding to the epitope bound by inhibitory anti-RAP-1 antibodies, and four other RAP-1 and RAP-2 synthetic peptides were used in these studies. Plasma from the native population had greater IgG reactivity to the N-terminal third of RAP-1 than the migrant population; both populations had low levels of IgM to this region of RAP-1. The IgG reactivity to RAP-2 and to the C-terminal third of RAP- 1, as well as for all the peptides, including the peptide from the inhibitory domain, were low or absent in both populations. In contrast, there were a high number of subjects with an IgM response to the peptides. Cellular responses were measured by proliferation of peripheral blood mononuclear cells (PBMC) and, in some subjects, by reverse transcription-polymerase chain reaction for interleukin-2 (IL- 2), interferon-gamma (IFN-gamma), IL-4, and IL-10. Proliferation of PBMC was low when stimulated by recombinant proteins, peptides, or parasite lysate. Both RAP-1 and RAP-2 stimulated cytokine production by donor T cells; IL-2, IL-4, and IFN-gamma RNA transcripts were observed in response to recombinant proteins and parasite lysate, but with no uniform trends. From the observed antibody responses, RAP-1 appears to be more immunogenic than RAP-2.Am J Trop Med HygT 1998592` 208-1693114883|Jakobsen, P. H. Hviid, L. Theander, T. G. Afare, E. A. Ridley, R. G. Heegaard, P. M. Stuber, D. Dalsgaard, K. Nkrumah, F. K.Specific T-cell recognition of the merozoite proteins rhoptry- associated protein 1 and erythrocyte-binding antigen 1 of Plasmodium falciparumAdolescence Adult Amino Acid Sequence Antigens, Surface/*immunology Carrier Proteins/*immunology Child Child, Preschool Enzyme-Linked Immunosorbent Assay Human IgG/biosynthesis IgM/biosynthesis *Immunity, Cellular Lymphocyte Transformation/immunology Malaria, Falciparum/*immunology Middle Age Molecular Sequence Data Protein Precursors/immunology Protozoan Proteins/*immunology Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. T-Lymphocytes/*immunologyvpThe merozoite proteins merozoite surface protein 1 (MSP-1) and rhoptry- associated protein 1 (RAP-1) and synthetic peptides containing sequences of MSP-1, RAP-1, and erythrocyte-binding antigen 1, induced in vitro proliferative responses of lymphocytes collected from Ghanaian blood donors living in an area with a high rate of transmission of malaria. Lymphocytes from a large proportion of the Ghanaian blood donors proliferated in response to the RAP-1 peptide, unlike those of Danish control blood donors, indicating that this sequence contains a malaria-specific T-cell epitope broadly recognized by individuals living in an area with a high transmission rate of malaria. Most of the donor plasma samples tested contained immunoglobulin G (IgG) and IgM antibodies recognizing the merozoite proteins, while only a minority showed high IgG reactivity to the synthetic peptides. Infect Immun 1993611 268-73>`2(hkGp2(Freeman, R. @2(GkG GGgkGkGkGkGG ؏GGGkGGkGkGGikGٳkGkGkGٳG ؏GGGkGkGkkGkGGkkG kGkGkGkGG GGG kGGGkGݳGk߳kG[GسkGkGk.'Etzion, Z. Murray, M. C. Perkins, M. E.JDIsolation and characterization of rhoptries of Plasmodium falciparum 1991Mol Biochem Parasitol471o 51-62$f`http://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://iai.asm.org/cgi/content/full/66/3/1037981477170)Fonjungo, P. N. Stuber, D. McBride, J. S.ngAntigenicity of recombinant proteins derived from rhoptry-associated protein 1 of Plasmodium falciparumajdAdolescence Adult Animal Antibodies, Protozoan/analysis/classification Antigens, Protozoan/*immunology Child Child, Preschool Enzyme-Linked Immunosorbent Assay Epitopes Female Human IgG/classification Immune Sera/immunology Infant Mice Plasmodium falciparum/*immunology Protozoan Proteins/*immunology Recombinant Proteins/immunology Support, Non-U.S. Gov'tRhoptry-associated protein 1 (RAP1) of Plasmodium falciparum is a potential component of a malaria vaccine. We have expressed in Escherichia coli eight recombinant RAP1 proteins representing almost the entire sequence of the mature protein and assessed the antigenicity of the proteins by immunization of mice. Antisera to six of the recombinant proteins reacted specifically with parasite-derived RAP1 (PfRAP1), as determined by indirect immunofluorescence and by immunoblotting. These proteins were then used in enzyme-linked immunosorbent assays to evaluate human antibody responses to RAP1 during naturally transmitted infections in The Gambia. Immunoglobulin G (IgG) antibodies specifically reactive with the recombinant RAP1 proteins are directed mostly towards fragments containing the N- terminal sequences of mature PfRAP1. The most N-terminal segment (residues 23 to 175) contains only minor epitopes, while major epitopes are outside this region. Antibodies from malaria patients do not compete for a linear epitope recognized by an inhibitory anti-RAP1 monoclonal antibody. Analysis of IgG subclass distribution shows that human anti-RAP1 antibodies are predominantly IgG1. Infect Immun 19986631037-44`T|EZ852371504-Stewart, M. J. Schulman, S. Vanderberg, J. P.uNHRhoptry secretion of membranous whorls by Plasmodium berghei sporozoitesAnimal Glutaral Intracellular Membranes/*ultrastructure Microscopy, Electron Movement Organoids/ultrastructure Plasmodium berghei/physiology/*ultrastructure Support, U.S. Gov't, P.H.S. Tannic Acid60Electron microscopy of sporozoites of the rodent malaria parasite, Plasmodium berghei, reveals electron-dense multilaminate membranous whorls within components of the rhoptry-microneme complex after fixation with tannic acid in conjunction with glutaraldehyde. This multilaminate material, which has a dark line to dark line periodicity of approximately 5 nm, appears to be secreted from the sporozoite since it is also found adhering to the sporozoite's external surface. The material may function in sporozoite gliding motility and in invasion of host cells. J Protozool0 1985322 280-396431681VOStowers, A. Prescott, N. Cooper, J. Takacs, B. Stueber, D. Kennedy, P. Saul, A.d^WImmunogenicity of recombinant Plasmodium falciparum rhoptry associated proteins 1 and 2AAmino Acid Sequence Animal Antigens, Protozoan/*immunology Disease Models, Animal Enzyme-Linked Immunosorbent Assay *Epitope Mapping Erythrocytes/cytology/immunology Female IgG/blood/immunology Immunoblotting Immunogenetics Malaria Vaccines/*immunology Mice Molecular Sequence Data Plasmodium falciparum/*immunology Protozoan Proteins/*immunology Rabbits Recombinant Fusion Proteins/immunology Support, Non-U.S. Gov'ts(!Mice and rabbits immunized with recombinant forms of malaria vaccine candidate antigens rhoptry-associated proteins 1 and 2 (RAP-1, RAP-2 and rRAP-1, rRAP-2) produce antibodies at titres equivalent to monoclonal antibody ascites fluid raised against the native proteins. Sera from animals immunized with rRAP-1 contain antibodies which recognize the native protein by indirect immunofluorescence and immunoblotting, partially inhibit erythrocyte invasion in vitro and are long lasting. Epitope mapping shows these antibodies predominantly recognize epitopes in the N-terminal third of rRAP-1, some of which coincide with the targets of inhibitory monoclonal antibodies. By contrast, sera from animals immunized with rRAP-2 contain antibodies which recognize the recombinant but not the native protein.oParasite Immunol 19951712 631-4298299938,%Stowers, A. W. Spring, K. J. Saul, A.b\Preparative scale purification of recombinant proteins to clinical grade by isotachophoresis`ZElectrophoresis/instrumentation/*methods Electrophoresis, Polyacrylamide Gel Malaria Vaccines/isolation & purification Protein Denaturation Protozoan Proteins/isolation & purification Recombinant Proteins/*isolation & purification Serum Albumin, Bovine/isolation & purification Support, Non-U.S. Gov't Vaccines, Synthetic/isolation & purification:4An electrophoretic procedure based on isotachophoresis has been developed for protein purification on a preparative scale in the 10 to 500 mg range. The system is simple, uses well understood physical properties, does not need ampholyte spacers and is able to produce sterile products of clinical grade. We demonstrate the applicability of this apparatus for the purification of denatured recombinant proteins and complex mixtures of proteins. The system may also be used for both cationic and anionic purification of proteins in their native form. The system is scalable from analytical to preparative protein loads at consistently high protein yields and purity levels. Total protein loads may vary as much as 1000 fold with the use of interchangeable columns of varying diameter and constant length. At both preparative and analytical scales concentration of products at greater than 20 mg/ml are obtainable. Toxicological considerations are addressed with assays for endotoxin, acrylamide and SDS concentrations, as well as the prevention of covalent protein modification.Biotechnology (N Y)e 19951313 1498-1503iMO P@(l, A.H%s (Jarra, W.P. (;Аc};c<|%`簀|:H>`,A99066929Noe, A. R. Adams, J. H.^WPlasmodium yoelii YM MAEBL protein is coexpressed and colocalizes with rhoptry proteinsH Animal Antibodies, Protozoan/immunology Carrier Proteins/*analysis/biosynthesis/chemistry/immunology Cell Membrane/chemistry Chimeric Proteins/chemistry/immunology Fluorescent Antibody Technique Membrane Proteins/analysis/immunology Merozoite Surface Protein 1/analysis Organelles/chemistry Plasmodium yoelii/*chemistry/growth & development/metabolism Precipitin Tests Protozoan Proteins/analysis/immunology Receptors, Cell Surface/*analysis/biosynthesis/chemistry/immunology Solubility Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.`YWe have previously cloned genes from multiple rodent malaria species exhibiting characteristics of the genes encoding Duffy binding like- erythrocyte binding proteins (DBL-EBP). Homology is seen in the intron/exon structure of the genes and in the carboxyl terminal region (including the deduced carboxyl cysteine-rich domain) of the proteins they encode. However, the amino termini of these proteins are not homologous to the DBL-EBP but contain tandem cysteine-rich regions that are similar to the cysteine-rich region of AMA-1 (apical membrane antigen-1), a rhoptry protein. This new family of proteins has been termed MAEBL and these are paralogues of both AMA-1 and the DBL-EBP. Serum against the carboxyl cysteine-rich region of the Plasmodium yoelii YM MAEBL reacted to parasites with a punctate fluorescence pattern characteristic of apical organelle proteins and also localized MAEBL to the surface of merozoites within schizonts. This antiserum immunoprecipitated a protein doublet (120/128 kDa) that was unexpectedly insoluble when compared to members of the DBL-EBP. Characterization of MAEBL was extended through colocalization studies comparing the P. yoelii YM MAEBL to other parasite proteins. This protein appeared to be located in the rhoptry organelles as it colocalized with both AMA-1 and the P. yoelii 235 kDa rhoptry proteins within parasites. In addition, MAEBL is expressed relatively early in schizont development and appears on the merozoite surface after segmentation. Both the pattern and time of expression of the P. yoelii YM MAEBL are consistent with a rhoptry rather than a microneme protein.GMol Biochem Parasitolc 199896 1-2h 27-35D>Owen, C. A. Sinha, K. A. Keen, J. K. Ogun, S. A. Holder, A. A. 1999^XChromosomal organisation of a gene family encoding rhoptry proteins in Plasmodium yoeliiMol Biochem Parasitol992 183-9299270308 The genomic organisation of the genes coding for a group of high molecular mass rhoptry proteins of the rodent malaria parasite Plasmodium yoelii YM was investigated using blotting, two dimensional gel electrophoresis and restriction fragment length analysis. The genes were found on chromosomes 1, 5, 6 and 10, with the possibility that related genes were also present on chromosomes 3 and 4. On chromosome 1 the genes were located close to one end, whereas they were present at both ends of chromosome 5, 6 and 10. Two genes, e3 and e8, that had been partially characterised previously were present on chromosomes 5 and 1, respectively. Based on an analysis of the 3' end of the genes, three subfamilies present on chromosomes 1, 5 and 6, and 10, respectively, were identified. 878:01L:pcroft, J. A. Kidson, C. (Kidson, C..   (GGGGGGG޳90348711voRidley, R. G. Takacs, B. Lahm, H. W. Delves, C. J. Goman, M. Certa, U. Matile, H. Woollett, G. R. Scaife, J. G.e^XCharacterisation and sequence of a protective rhoptry antigen from Plasmodium falciparumpiAmino Acid Sequence Animal *Antigens, Protozoan/genetics/immunology/isolation & purification Base Sequence Blotting, Western Cloning, Molecular Electrophoresis, Polyacrylamide Gel Genes Molecular Sequence Data Plasmodium falciparum/genetics/*immunology *Protozoan Proteins/genetics/immunology/isolation & purification Restriction Mapping Support, Non-U.S. Gov'taWe have recently demonstrated that a non-polymorphic rhoptry antigen, RAP-1 (rhoptry associated protein-1), which is recognised by human immune serum, can successfully protect Saimiri monkeys from a lethal infection of Plasmodium falciparum malaria. In this report we further characterise the antigen, which consists of four major proteins of 80, 65, 42 and 40 kDa and two minor proteins of 77 and 70 kDa, and present the antigen's gene sequence. Monoclonal antibody evidence, autocatalytic processing and immunological cross-reactivity suggest that all components of this antigen are derived from the same precursor protein. The antigen is lipophilic, and disulphide bonding plays an important role in its structure. We discuss the structure and function of RAP-1 in the light of its deduced amino acid sequence and consider the relationship of this antigen to other rhoptry antigens of similar subunit size and composition.Mol Biochem Parasitol  1990411 125-3482Ridley, R. G. Lahm, H. W. Takcs, B. Scaife, J. G.zGenetic and structural relationships between components of a protective rhoptry antigen complex from Plasmodium falciparum 1991Mol Biochem Parasitolc472245-246 Ridley, R. G..`YProteins of unusual sequence composition from the malarial parasite Plasmodium falciparum 1991Biochem Soc Trans192525-528RKRoger, N. Dubremetz, J. F. Delplace, P. Fortier, B. Tronchin, G. Vernes, A. 1988TMCharacterization of a 225 kilodalton rhoptry protein of Plasmodium falciparumMol Biochem Parasitol27135-141| 0634 Ref11+.(Sam-Yellowe, T.Y. Shio, H. Perkins, M.E.f`Secretion of Plasmodium falciparum rhoptry protein into the plasma membrane of host erythrocytesJ Cell Biology 1988 106] 1507-1513@ Ref11n& Sam-Yellowe, T. Y. Perkins, M.E. 1990lBinding of Plasmodium falciparum rhoptry proteins to mouse erythrocytes and their possible role in invasion.     Mol Biochem Parasitolu392n 91-100 0068 Monash&Sam-Yellowe, T.Y. Perkins, M.E. 1991Interaction of the 140/130/110 kDa rhoptry protein complex of Plasmodium falciparum with the erythrocyte membrane and liposomes > S Exp. Parasitol.73161-171 1436 Ref11Sam-Yellowe, T.Y. 1992d]Molecular factors responsible for host cell recognition and invasion in Plasmodium falciparum J Protozool391 181-9Sam-Yellowe, T. Y. 1993Plasmodium falciparum: Analysis of protein-protein interactions of the 140/130/110-kDa rhoptry protein complex using antibody and mouse erythrocyte binding assays Exp Parasitol 772 179-194 286 Monash? D8W  Howard, R. F. 1992lfThe sequence of the p82 rhoptry protein is highly conserved between two Plasmodium falciparum isolatesMol Biochem Parasitol51 327932933172+Howard, R. F. Jensen, J. B. Franklin, H. L.eReactivity profile of human anti-82-kilodalton rhoptry protein antibodies generated during natural infection with Plasmodium falciparum0)Amino Acid Sequence Animal Antibodies, Protozoan/*immunology Base Sequence Epitopes Human Malaria, Falciparum/*immunology Molecular Sequence Data Plasmodium falciparum/*immunology Protozoan Proteins/*immunology Repetitive Sequences, Nucleic Acid Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.NRKImmunization of monkeys with the 82-kDa rhoptry protein (p82) of Plasmodium falciparum can protect them against a lethal blood stage challenge, and monoclonal antibodies to p82 inhibit parasite growth in vitro. The role that a p82-specific immune response might play in human immunity to the parasite is not known. To determine to what extent humans produce antibodies to p82 following infection with P. falciparum, sera from individuals believed to be hyperimmune, semi- immune, or never infected with the parasite were examined. Portions of the p82 gene were expressed separately as fusion proteins and used on immunoblots to test for antibodies to the recombinant proteins. All but 1 of the 30 immune sera possessed antibodies to p82, while nonimmune sera produced, at best, only a marginal signal to the fusion proteins. The signal intensity produced with the human immune sera depended on the region of p82 being assayed, with the N-terminal 37% of p82 producing stronger signals than more C-terminal parts of p82. This N- terminal domain contains a tandem octapeptide repeat (consensus KSSSPSXT/V) of the structure (repeat)2-Q-T-S-G-S/L-(repeat)3. It is shown here that the sequence of this repetitive motif is conserved among four parasite isolates at both the nucleotide and amino acid levels; the five-residue repeat interruption peptide QTSGS/L separating the two sets of repeats contains the only amino acid substitution (Ser or Leu) detected in this region to date. Despite their conservation of structure, the repeats do not appear to be the only epitope recognized by the human antibodies, since sera which recognize the N-terminal fusion protein containing the repeats also bind a related protein after truncation and removal of the repeats. These results indicate that the structurally conserved p82 molecule contains multiple B-cell epitopes and is likely to be immunogenic in most individuals during natural infections with P. falciparum. These observations are consistent with the idea that antibodies to p82 generated during parasite infection have a role in the development of immunity to the organism.t Infect Immun 1993617 2960-596379224 Howard, R. F. Peterson, C.RKLimited RAP-1 sequence diversity in field isolates of Plasmodium falciparumFvpAmino Acid Sequence Animal Antigens, Protozoan/chemistry/*genetics Base Sequence Codon Comparative Study Genes, Protozoan Geography Human Malaria, Falciparum/blood Molecular Sequence Data Plasmodium falciparum/*genetics/isolation & purification/metabolism Protozoan Proteins/chemistry/*genetics Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. *Variation (Genetics)Mol Biochem Parasitol  1996771 95-8f_http://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://iai.asm.org/cgi/content/full/66/1/38098084502:4Howard, R. F. Jacobson, K. C. Rickel, E. Thurman, J.Analysis of inhibitory epitopes in the Plasmodium falciparum rhoptry protein RAP-1 including identification of a second inhibitory epitopengAmino Acid Sequence Animal Antibodies, Monoclonal/biosynthesis/immunology Antibodies, Protozoan/*analysis Aotinae B-Lymphocytes/immunology Cells, Cultured Cloning, Molecular Epitopes/*immunology Erythrocytes/parasitology Immunity, Active Immunodominant Epitopes Malaria, Falciparum/blood/*immunology Male Mice Molecular Sequence Data Parasitemia/immunology Plasmodium falciparum/genetics/*immunology Protozoan Proteins/genetics/*immunology Recombinant Fusion Proteins/immunology Recombination, Genetic Sequence Homology, Amino Acid Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.o Immune responses to Plasmodium falciparum rhoptry-associated protein 1 (RAP-1), RAP-2, and RAP-3 appear to contribute to protection against infection by this human malarial parasite. This conclusion is suggested by results of monkey immunization trials and of cell culture studies showing antibody-dependent inhibition of erythrocyte invasion. In the present study, splenectomized owl monkeys were infected with P. falciparum in order to monitor anti-RAP-1 antibody production as antiparasite immunity developed. The monkeys responded to a primary infection with the production of antibodies to a fragment of RAP-1 containing amino acids 1 to 294 (RAP-1(1-294)). After drug cure and reinfection, the monkeys had a prolonged prepatent period, indicating they had already developed partial immunity to the parasite. Sera from these animals showed major increases in anti-RAP-1(1-294) antibodies. In contrast, only low levels of antibodies to inhibitory B-cell epitope 1 (iB-1), an inhibitory epitope in RAP-1(1-294) with the sequence N200TLTPLEELYPT211, was observed after the initial parasite infection, and the anti-iB-1 antibodies were not readily boosted upon reinfection. These results suggest that iB-1 is an immunogenic but not immunodominant epitope and that anti-iB-1 antibodies do not substantially contribute to early stages of naturally acquired immunity in the owl monkey model. To identify additional epitopes bound by inhibitory antibodies, mouse monoclonal antibodies were produced with a recombinant fusion protein containing RAP-1(1-294). Monoclonal antibody 1D6 inhibited parasite invasion of erythrocytes in vitro. 1D6 did not bind peptide iB-1 but rather bound a second inhibitory epitope called iB-2. iB-2, like iB-1, is found near the amino terminus of p67, a RAP-1 processing product thought to be involved in merozoite invasion of erythrocytes. Since anti-iB-1 antibodies were not readily produced during parasite infection, it may be desirable to direct antibody responses to particular epitopes in RAP-1, such as iB-1 and iB-2. Infect Immun 1998661 380-6 &J<[X%\PZN 6f`http://www.ncbi.nlm.nih.gov/cgi-bin/Entrez/referer?http://iai.asm.org/cgi/content/full/67/6/297599270961voFonjungo, P. N. Elhassan, I. M. Cavanagh, D. R. Theander, T. G. Hviid, L. Roper, C. Arnot, D. E. McBride, J. S. A longitudinal study of human antibody responses to Plasmodium falciparum rhoptry-associated protein 1 in a region of seasonal and unstable malaria transmissionvoAdult Animal Antibodies, Protozoan/blood/*immunology Antigens, Protozoan/*immunology Cohort Studies Human IgG/blood/immunology IgM/blood/immunology Longitudinal Studies Malaria, Falciparum/blood/*immunology/transmission Plasmodium falciparum/immunology Protozoan Proteins/*immunology Recombinant Fusion Proteins/immunology Seasons Support, Non-U.S. Gov't Time Factors/Rhoptry-associated protein 1 (RAP1) of Plasmodium falciparum is a nonpolymorphic merozoite antigen that is considered a potential candidate for a malaria vaccine against asexual blood stages. In this longitudinal study, recombinant RAP1 (rRAP1) proteins with antigenicity similar to that of P. falciparum-derived RAP1 were used to analyze antibody responses to RAP1 over a period of 4 years (1991 to 1995) of 53 individuals naturally exposed to P. falciparum malaria. In any 1 year during the study, between 23 and 39% of individuals who had malaria developed immunoglobulin G (IgG) antibodies detectable with at least one rRAP1 protein. However, the anti-RAP1 antibody responses were detected only during or shortly after clinical malarial infections. RAP1 antibody levels declined rapidly (within 1 to 2 months) following drug treatment of the infections. No anti-RAP1 antibodies were usually detected a few months after the end of malaria transmission, during the dry season, or by the start of the next malaria season. Thus, RAP1 IgG responses were very short-lived. The short duration of RAP1 antibody response may explain the apparent lack of response in a surprisingly high proportion of individuals after clinical malarial infections. For some individuals who experienced more than one malarial infection, a higher anti-RAP1 antibody response to subsequent infections than to earlier infections was observed. This suggested secondary responses to RAP1 and thus the development of immunological memory for RAP1. Infect Immun 19996762975-85tTMHarnyuttanakorn, P. McBride, J. J. Donachie, S. Heidrich, H. G. Ridley, R. G. 1992Inhibitory monoclonal antibodies recognise epitopes adjacent to a proteolytic cleavage site on the RAP-1 protein of Plasmodium falciparum t Mol. Biochem. Parasitol.55177-186d Rap-1VPHienne, R. Ricard, G. Fusa, T. Fujioka, H. Pradines, B. Aikawa, M. Doury, J.-C. 1998QPlasmodium yoelii: Identification of rhoptry proteins using monoclonal antibodiesm  Exp. Parasitol.903230-235 Experimental Parasitologyg Py MalAg MalMab"Holder, A. A. Freeman, R. R.VPImmunization against blood-stage rodent malaria using purified parasite antigens Nature 1981 294\361-364 Ref1185113444"Holder, A. A. Freeman, R. R.@:Protective antigens of rodent and human bloodstage malaria@:Animal Antibodies, Monoclonal Antigens, Protozoan/*immunology/isolation & purification Comparative Study Human Malaria/blood/*immunology/prevention & control Membrane Proteins/immunology Molecular Weight Plasmodium/growth & development/*immunology Plasmodium falciparum/immunology Vaccines/isolation & purificationBloodstage malaria parasites are antigenically complex, but individual antigens can be identified and analysed using monoclonal antibodies. Two monoclonal antibodies that recognize a 235 000 molecular mass Plasmodium yoelii rhoptry protein provide some protection when injected into mice against a challenge infection. The purified rhoptry protein also provides protective immunity against P. yoelii YM when used to vaccinate mice and fulminating infections are converted into self- limiting, reticulocyte-restricted infections. Another monoclonal antibody immunoprecipitates a 230 000 molecular mass protein and a series of proteolytic processing fragments. At least one of these processing fragments, probably a 90 000 molecular mass species, is located on the merozoite surface. Mice immunized with the purified protein were protected against challenge infection with P. yoelii YM. This antigen may provide protection by inducing a cell-mediated immune response. A monoclonal antibody raised against P. falciparum schizonts reacts with a 195 000 molecular mass protein which is synthesized in schizonts and subsequently cleaved. Fragments of the 195 000 molecular mass protein are expressed as major antigens on the merozoite surface. The 195 000 molecular mass P. falciparum protein and the 230 000 molecular mass P. yoelii protein belong to a class of malaria parasite antigens which probably is important in the induction of a protective immune response in the host.("Philos Trans R Soc Lond B Biol Sci 1984 307  1131 171-7851880676/Holder, A. A. Freeman, R. R. Uni, S. Aikawa, M.a:4Isolation of a Plasmodium falciparum rhoptry proteinAnimal Antibodies, Monoclonal Antigens, Protozoan/immunology/*isolation & purification Comparative Study Histocytochemistry Molecular Weight Organoids/analysis Plasmodium falciparum/*analysis/ultrastructure Support, Non-U.S. Gov'tA monoclonal antibody raised against the malaria parasite Plasmodium falciparum recognised a protein of 140000 molecular weight which was synthesized during schizogony. The protein has been purified by monoclonal antibody affinity chromatography from extracts of parasitized red cells. Antibodies against the protein have been used to determine its subcellular location. The protein is not expressed on the merozoite surface and has been located in the rhoptries, the apical organelles of the merozoite.sMol Biochem Parasitoli 1985143293-30392320857:3Holder, A. A. Keen, J. K. Sinha, K. A. Brown, K. N.6/The 235 kD rhoptry protein of Plasmodium yoelii.Amino Acid Sequence Animal Antigens, Protozoan/*genetics/immunology Base Sequence Molecular Sequence Data Multigene Family Plasmodium yoelii/*geneticsA 235 kD rhoptry protein produced by the malaria parasite, Plasmodium yoelii is the target of antibodies that protect mice against blood- stage challenge with the virulent YM strain. In the protected animals the parasites are confined to reticulocytes and the course of parasitaemia is reminiscent of an avirulent 17X strain infection. The DNA coding for the rhoptry protein has been identified as a multigene family containing at least four members. Sequence analysis of short DNA clones has identified the C-terminus of the protein; a preliminary analysis of longer clones confirms that the genes are polymorphic. The possible implications of these findings for the biology of the parasite are discussed. 1991 Acta Leiden601 101-6 Using Smart Source Parsing<5Howard, R.F. Stanley, H.A. Campbell, G.H. Reese, R.T. 1984b\Proteins responsible for a punctate fluorescence pattern in Plasmodium falciparum merozoitesAm J Trop Med Hyg33 1055-1059 0146 Ref11 Howard, R. F. Reese, R. T.RKPlasmodium falciparum: Hetero-oligomeric complexes of rhoptry polypeptides Exp Parasitol 1990713330-3426/Reference Number: 1458; Reference Type: Articlen Howard, R.F. 1990{The lower-molecular-weight protein complex (RI) of the Plasmodium falciparum rhoptries lacks the glycolytic enzyme aldolaseMol Biochem ParasitolP42235-240 0986 Ref11   Aikawa, M. Blackman, M. Collins, W.E. Deloron, P. Doury, J.-C. Dubois, B. Fievet, N. Fujioka, H. Fusa, T. Hasnain, S.E. Hienne, R. Hoffman, S.L.Holloway, B.P. Howard, R.F. Kumar, N. Lal, A.A. Luty, A.J.F. Mayombo, J. Mbessi, J.R.Migot-Nabias, F. Millet, P. Minh, T.N. Narum, D.L. Pradines, B. Renaut, A. Ricard, G. Ringwald, P. Sacci, J.B. Shi, Y.P. Thurman, J Vaillant, M.Wohlhueter, R. (LHAm. J. Trop. Med. Hyg. American Journal of Tropical Medicine and Hygiene,)Exp. Parasitol. Experimental ParasitologyD?Mol. Biochem. Parasitol. Molecular and Biochemical ParasitologyliProc. Nat. Acad. Sci. USA Proceedings of the National Academy of Sciences of the United States of AmericaMO P^Migot-Nabias, F. Luty, A.J.F. Ringwald, P. Vaillant, M. Dubois, B. Renaut, A. Mayombo, J. Minh, T.N. Fievet, N. Mbessi, J.R. Millet, P. Deloron, P. 1999Immune responses against Plasmodium falciparum asexual blood-stage antigens and disease susceptibility in Gabonese and Cameroonian children  . Am. J. Trop. Med. Hyg.613.488-49481American Journal of Tropical Medicine and Hygiene99066929Noe, A. R. Adams, J. H.^WPlasmodium yoelii YM MAEBL protein is coexpressed and colocalizes with rhoptry proteinsH Animal Antibodies, Protozoan/immunology Carrier Proteins/*analysis/biosynthesis/chemistry/immunology Cell Membrane/chemistry Chimeric Proteins/chemistry/immunology Fluorescent Antibody Technique Membrane Proteins/analysis/immunology Merozoite Surface Protein 1/analysis Organelles/chemistry Plasmodium yoelii/*chemistry/growth & development/metabolism Precipitin Tests Protozoan Proteins/analysis/immunology Receptors, Cell Surface/*analysis/biosynthesis/chemistry/immunology Solubility Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.`YWe have previously cloned genes from multiple rodent malaria species exhibiting characteristics of the genes encoding Duffy binding like- erythrocyte binding proteins (DBL-EBP). Homology is seen in the intron/exon structure of the genes and in the carboxyl terminal region (including the deduced carboxyl cysteine-rich domain) of the proteins they encode. However, the amino termini of these proteins are not homologous to the DBL-EBP but contain tandem cysteine-rich regions that are similar to the cysteine-rich region of AMA-1 (apical membrane antigen-1), a rhoptry protein. This new family of proteins has been termed MAEBL and these are paralogues of both AMA-1 and the DBL-EBP. Serum against the carboxyl cysteine-rich region of the Plasmodium yoelii YM MAEBL reacted to parasites with a punctate fluorescence pattern characteristic of apical organelle proteins and also localized MAEBL to the surface of merozoites within schizonts. This antiserum immunoprecipitated a protein doublet (120/128 kDa) that was unexpectedly insoluble when compared to members of the DBL-EBP. Characterization of MAEBL was extended through colocalization studies comparing the P. yoelii YM MAEBL to other parasite proteins. This protein appeared to be located in the rhoptry organelles as it colocalized with both AMA-1 and the P. yoelii 235 kDa rhoptry proteins within parasites. In addition, MAEBL is expressed relatively early in schizont development and appears on the merozoite surface after segmentation. Both the pattern and time of expression of the P. yoelii YM MAEBL are consistent with a rhoptry rather than a microneme protein.GMol Biochem Parasitolc 199896 1-2h 27-35D>Owen, C. A. Sinha, K. A. Keen, J. K. Ogun, S. A. Holder, A. A. 1999^XChromosomal organisation of a gene family encoding rhoptry proteins in Plasmodium yoeliiMol Biochem Parasitol992 183-9299270308 The genomic organisation of the genes coding for a group of high molecular mass rhoptry proteins of the rodent malaria parasite Plasmodium yoelii YM was investigated using blotting, two dimensional gel electrophoresis and restriction fragment length analysis. The genes were found on chromosomes 1, 5, 6 and 10, with the possibility that related genes were also present on chromosomes 3 and 4. On chromosome 1 the genes were located close to one end, whereas they were present at both ends of chromosome 5, 6 and 10. Two genes, e3 and e8, that had been partially characterised previously were present on chromosomes 5 and 1, respectively. Based on an analysis of the 3' end of the genes, three subfamilies present on chromosomes 1, 5 and 6, and 10, respectively, were identified.