Apical expression of three RhopH1/Clag proteins as components of the Plasmodium falciparum RhopH complex

S eventeen million people die of transmittable diseases and 2/3 of the world's population suffer them annually. Malaria, tuberculosis, AIDS, hepatitis, and reemerging and new diseases are a great threat to humankind. A logical and rational approach for vaccine development is thus desperately needed.Protein chemistry provides the best tools for tackling these problems. The tremendous complexity of microbes, the different pathways they use for invading host cells, and the immune responses they induce can only be resolved by using the minimum subunit-based (chemically produced ∼20-mer peptides), multiantigenic (most proteins involved in invasion), multistage (different invasion mechanisms) vaccine development approach.The most lethal form of malaria caused by Plasmodium falciparum (killing 3 million and affecting 500 million people worldwide annually) was used as target disease since many of its proteins, its invasion pathways, and its genome have been described recently. A New World primate (the Aotus monkey) is highly susceptibly to human malaria; its immune system molecules are 80 -100% identical to those of its human counterpart, making it an excellent model for vaccine development.Chemically synthesized ∼20-mer peptides, covering all the P. falciparum malaria proteins involved in red blood cell (RBC) invasion were synthesized by the classical t-Boc technology (based on synthetic SPf66 antimalarial vaccine information for identifying targets) and assayed in a highly sensitive, specific, and robust test for detecting receptor-ligand interactions between high-activity binding peptides (HABPs) and RBCs. HABPs were identified, some in which the molecule displays genetic variability (to be discarded due to their tremendous complexity) and elicits a strain-specific immune response and others that are conserved (no amino acid sequence variation).Conserved HABPs were synthesized in a polymeric form by adding cysteines at their N-and C-terminal ends to be used for monkey immunization. They became nonimmunogenic (no antibodies were induced) nonprotection inducers (monkeys were not protected against P. falciparum malaria challenge with a highly infective strain) suggesting a code of immunological silence or nonresponsiveness for these conserved HABPs.A large number of monkey trials involving a considerable number of Aotus monkeys were performed to break this code of immunological silence by replacing critical residues (determined by glycine peptide analogue scanning) to find that the following amino acid changes had to be made to render them antibody and protection inducing: FTR; WTY; LTH; ITN; MTK; PTD; QTE; CTT.The three-dimensional (3D) structure of >100 of these native modified HABPs (determined by 1 H NMR) revealed that the following structural changes had all to be achieved to allow a better fit into the major histocompatibility complex class II (MHC II)-peptide-TCR complex to properly activate the immune system: R-helix shortening, modifying their -turn, adopting segmental R-helix configuration, changing residue or...

show abstract

“…CLAG9, and the CLAG family (CLAG-1-9) 17 [ Figure 1B]. However, their role has still to be fully established.…”

Section: Rationale For Multiantigenic Multistage Vaccine Developmentmentioning

confidence: 99%

Emerging Rules for Subunit-Based, Multiantigenic, Multistage Chemically Synthesized Vaccines

2008

View full text Add to dashboard Cite

show abstract

“…The genes encoding RhopH1 are members of the rhoph1/clag gene family, which was originally defined by the cytoadherence linked asexual gene (clag) on chromosome 9 in P. falciparum (clag9) and consists of at least three members; clag2, 3.1, and 9 [13][14][15].…”

Section: Introductionmentioning

confidence: 99%

“…Although not yet determined experimentally, molecules encoded by clag3.2 and 8 are likely parts of the RhopH complex as judged by their similarity in amino acid sequence and transcription pattern with other members [15]. Because only one RhopH1/Clag participates to form a single RhopH complex [15,16], five types of PfRhopH complex are expected to exist, each of which contains one rhoph1/clag gene product.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Diversity and evolution of the rhoph1/clag multigene family of Plasmodium falciparum

Iriko

Kaneko

Otsuki

et al. 2008

Molecular and Biochemical Parasitology

Self Cite

View full text Add to dashboard Cite

A complex of high molecular mass proteins (PfRhopH) of the human malaria parasite Plasmodium falciparum induces host protective immunity and therefore is a candidate for vaccine development. Understanding the level of polymorphism and the evolutionary processes is important for advancements in both vaccine design and knowledge of the evolution of cell invasion in this parasite. In the present study, we sequenced the entire open reading frames of seven genes encoding the proteins of the PfRhopH complex (rhoph2, rhoph3, and five rhoph1/clag gene paralogs). We found that four rhoph1/clag genes (clag2, 3.1, 3.2, and 8) were highly polymorphic. Amino acid substitutions and indels are predominantly clustered around amino acid positions 1000-1200 of these four rhoph1/clag genes. An excess of nonsynonymous substitutions over synonymous substitutions was detected for clag8 and 9, indicating positive selection. The McDonald-Kreitman test with a Plasmodium reichenowi orthologous sequence also supports positive selection on clag8. Based on the ratio of interspecific genetic distance to intraspecific distance, the time to the most recent common ancestor of the clag2 and 8 polymorphisms was estimated to be 1.89 and 0.87 million years ago, respectively, assuming divergence of P. falciparum and P. reichenowi 6 million years ago. In addition to a copy number polymorphism, gene conversion events were detected for the rhoph1/clag genes on chromosome 3, which likely play a role in increasing the diversity of each locus. Our results indicate that a high diversity of the PfRhopH1/Clag multigene family is maintained by diversifying selection forces over a considerably long period.

show abstract

“…The PfRhopH complex containing CLAG proteins is composed of three subunits named RhopH1, RhopH2 and RhopH3 (Kaneko et al 2005). In the mature schizont, the subunits are localised in the merozoites' rhoptries, whose contents are discharged at the moment of contact with the erythrocyte membrane, concomitantly with the formation of the moving junction and the parasitophorous vacuole (PV).…”

mentioning

confidence: 99%

Cross-reactive anti-PfCLAG9 antibodies in the sera of asymptomatic parasite carriers of Plasmodium vivax

Costa

Zanchi

Rodrigues³

et al. 2013

Mem. Inst. Oswaldo Cruz

View full text Add to dashboard Cite

Apical expression of three RhopH1/Clag proteins as components of the Plasmodium falciparum RhopH complex

Cited by 75 publications

References 42 publications

Emerging Rules for Subunit-Based, Multiantigenic, Multistage Chemically Synthesized Vaccines

Emerging Rules for Subunit-Based, Multiantigenic, Multistage Chemically Synthesized Vaccines

Diversity and evolution of the rhoph1/clag multigene family of Plasmodium falciparum

Cross-reactive anti-PfCLAG9 antibodies in the sera of asymptomatic parasite carriers of Plasmodium vivax

Contact Info

Product

Resources

About