Biochemistry Z 0250 Intimate Molecular Interactions of P. falciparum Merozoite Proteins Involved in Invasion of Red Blood Cells and Their Implications for Vaccine Design -[443 refs.]. -(RODRIGUEZ, L. E.; CURTIDOR, H.; URQUIZA, M.; CIFUENTES, G.; REYES, C.; PATARROYO*, M. E.; Chem. Rev. (Washington, D. C.) 108 (2008) 9, 3656-3705; Inst. Immunol., Univ. Nac. Colombia, Bogota, Colombia; Eng.) -Lindner 50-274

in 1991. The principal focus of his research at the Fundación Instituto de Inmunología de Colombia is the molecular characterization of receptor-ligand interactions between pathogenic proteins involved in invasion and their corresponding host cell receptors, mainly for P. falciparum and M. tuberculosis. He is currently candidate for a Dr. Sc. degree in chemistry at the Universidad Nacional de Colombia.Mauricio Urquiza graduated in chemistry at the Universidad Nacional de Colombia in 1991. He is currently working as a researcher in the Virology department at the Fundación Instituto de Inmunología de Colombia and is candidate for a Dr. Sc. degree in chemistry at the Universidad Nacional de Colombia. The main focus of his research is the study of the host-parasite interactions, using as model P. falciparum, P. vivax, Human Papilloma virus, and Epstein-Barr virus. He is also working in the development of a Human Papilloma virus detection test.Gladys Cifuentes graduated in chemistry at the Universidad Nacional de Colombia in 1994 and is a member of the Nuclear Magnetic Resonance and Molecular Design section of the Three-Dimensional Structure Department at the Fundación Instituto de Inmunología de Colombia. Her research interests include 3D-structure and molecular design of peptides and proteins with shown biological and chemical relevance in host-parasite interactions.

The conserved, nonantigenic, nonimmunogenic malaria Merozoite Surface Protein-2 peptide 1, having high affinity for red blood cells, was rendered immunogenic and protective in Aotus monkeys by specifically changing some critical residues. The NMR structure revealed a switch from classical type III' into distorted III' and III beta turns in the protective peptides. These changes may lead to a better fit into the Aotus MHC class II human HLA-DRbeta1 12 molecule equivalent, thus activating the immune system.

Developing a rational methodology for obtaining vaccines against P. falciparum malaria (the disease's most lethal form, afflicting more than 250 million people around the world per year and killing about 2 million of them) [1] has become one of the main objectives of public health authorities around the world. [2] A multiantigenic vaccine, containing molecules from the parasite's different developmental stages, is required due to the parasite's remarkable complexity and adaptability. [3] The first such approach (the SPf66 synthetic vaccine), [4,5] which used peptides from molecules from different parasite stages, conferred limited protective efficacy in Aotus monkey studies and in field trials carried out on human volunteers around the world. [6] [a] Prof.

As microbes use many mechanisms for avoiding immunological pressure, new strategies must be developed to bypass the immunological code of silence of conserved, functionally-important amino acid sequences, such as those involved in high activity binding peptides' (HABPs) attaching to their host cells. Hundreds of experiments in large numbers of Aotus monkeys revealed that this immunological code of silence could be broken by shifting the polarity of some critical host cell binding residues in these HABPs by substituting F for R and vice versa, Y<-->W, L<-->H, I<-->N, P<-->D, M<-->K or E, C<-->T, V<-->N or S; there are special rules for A, G and S. (1)H-nuclear magnetic resonance of these modified, immunogenic, protection-inducing HABPs and molecular modelling revealed that such modifications induced appropriate fitting into specific HLA-DRbeta1 Pockets, suggesting the presence of new pockets and a haplotype- and allele-specific conscious TCR. A highly immunogenic and protection-inducing anti-malarial vaccine can thus be produced.

An anti-malarial vaccine against the extremely lethal Plasmodium falciparum is desperately needed. Peptides from this parasite's proteins involved in invasion and having high red blood cell-binding ability were identified; these conserved peptides were not immun genic or protection-inducing when used for immunizing Aotus monkeys. Modifying some critical binding residues in these high-activi binding peptides' (HABPs') attachment to red blood cells (RBC) allowed them to induce immunogenicity and protection against expermental challenge and acquire the ability to bind to specific HLA-DRp1* alleles. These modified HABPs adopted certain characterist structural configurations as determined by circular dichroism (CD) and 1H nuclear magnetic resonance (NMR) associated with certain HLA-DRβ1* haplotype binding activities and characteristics, such as a 2-Å-distance difference between amino acids fitting into HLA-DRp1 Pockets 1 to 9, residues participating in binding to HLA-DR pockets and residues making contact with the TCR, suggesting haplotyp and allele-conscious TCR. This has been demonstrated in HLA-DR-like genotyped monkeys and provides the basis for designing high effective, subunit-based, multi-antigen, multi-stage, synthetic vaccines, for immediate human use, malaria being one of them.

EBA-175 protein is used as a ligand in the binding of P. falciparum to red blood cells (RBCs). Evidence shows that the conserved peptide 1779 from this protein (with high red blood cell binding ability and known critical erythrocyte binding residues) plays an important role in the invasion process. This peptide is neither immunogenic nor protective; analogs having critical residues replaced by amino acids with similar volume or mass but different polarity were synthesized and inoculated into Aotus monkeys, and elicited different immunogenic and protective responses. Nuclear Magnetic Resonance (1H-NMR) studies revealed that peptide analog 21696 (non-immunogenic and non-protective) presents a large helical fragment, that the peptide 14012 (immunogenic and non-protective) helical fragment is smaller, while the peptide 22812 (immunogenic and protective) alpha-helix is shorter in a different region and possesses greater flexibility at its N-terminus. The presence of methionine residues could affect the structural stability of peptide 22812 and ultimately its immunological response. Our results suggest a new strategy for designing a new malaria multi-component subunit-based vaccine.

The erythrocyte binding antigen EBA-175 is a 175-kDa Plasmodium falciparum protein, which has been shown to be involved in the process of invasion of erythrocytes. It has been found that conserved peptide 1818 belonging to this protein has high red blood cell binding capacity and plays an important role in the invasion process. This peptide is neither immunogenic nor protective. Peptide 1818 analogues had some of their previously recognized critical red blood cell binding residues substituted for amino acids having similar volume or mass but different polarity to make them fit into HLA-DRbeta(1)*1101 molecules; these 1818 peptide analogues were then synthesized and inoculated into Aotus nancymaae monkeys, generating different immunogenic and/or protective immune responses. Short structures such as 3(10)-helix, classical, or distorted type-III beta-turns were found in the immunogenic and protective peptides once the secondary structure had been analyzed by NMR and its structure correlated with its immunological properties. These data suggest that peptide flexibility may lead to better fitting into immune system molecules, therefore making them excellent candidates for consideration as components of a subunit-based, multicomponent synthetic antimalarial vaccine.