Plasmodium falciparum rhoptry neck protein 4 has conserved regions mediating interactions with receptors on human erythrocytes and hepatocyte membrane.

Plasmodium falciparum-related malaria represents a serious worldwide public health problem due to its high mortality rates. P. falciparum expresses rhoptry neck protein 4 (PfRON4) in merozoite and sporozoite rhoptries, it participates in tight junction-TJ formation via the AMA-1/RON complex and is refractory to complete genetic deletion. Despite this, which PfRON4 key regions interact with host cells remain unknown; such information would be useful for combating falciparum malaria. Thirty-two RON4 conserved region-derived peptides were chemically synthesised for determining and characterising PfRON4 regions having high host cell binding affinity (high activity binding peptides or HABPs). Receptor-ligand interaction/binding assays determined their specific binding capability, the nature of their receptors and their ability to inhibit in vitro parasite invasion. Peptides 42477, 42479, 42480, 42505 and 42513 had greater than 2% erythrocyte binding activity, whilst peptides 42477 and 42480 specifically bound to HepG2 membrane, both of them having micromolar and submicromolar range dissociation constants (Kd). Cell-peptide interaction was sensitive to treating erythrocytes with trypsin and/or chymotrypsin and HepG2 with heparinase I and chondroitinase ABC, suggesting protein-type (erythrocyte) and heparin and/or chondroitin sulphate proteoglycan receptors (HepG2) for PfRON4. Erythrocyte invasion inhibition assays confirmed HABPs' importance during merozoite invasion. PfRON4 800-819 (42477) and 860-879 (42480) regions specifically interacted with host cells, thereby supporting their inclusion in a subunit-based, multi-antigen, multistage anti-malarial vaccine.

Peptide-pulsed dendritic cells' immunomodulating effect regarding Mycobacterium tuberculosis growth in macrophages.

Mycobacterium tuberculosis is one of the most successful pathogens affecting humans, being the main cause of tuberculosis. It accounts for most infectious agent-related deaths worldwide; it has been estimated that a third of the world's population are bacillus carriers. This pathogen's evolutionary adaptation is mainly due to its ability to block a host's immune system by preventing it using an effective immune response in cases of active tuberculosis. Peptide-based synthetic vaccines represent an alternative for counteracting tuberculosis; however, although peptide antigens can be identified, they are not recognised by a host's immune system. An approach using dendritic cells as immunomodulating agents for increasing synthetic peptides' antigenic capacity has thus been advanced. Dendritic cells obtained from IL to 4- and GM-CSF-treated peripheral blood mononuclear cells were pulsed with synthetic Mtb protein peptides which have been reported as participating in mycobacteria-host interactions; their amino acid sequences were modified to improve MHC-II coupling and thus increase their recognition by a host's immune system. pMHC-II/TCR interaction triggered a lymphocyte response which controlled Mtb intracellular growth in infected macrophages. This work has been aimed at contributing to understanding dendritic cells' role in Mycobacterium tuberculosis protein peptide antigen presentation, thereby increasing individuals' immune response as a means of controlling the disease.

Stereo electronic principles for selecting fully-protective, chemically-synthesised malaria vaccines.

Major histocompatibility class II molecule-peptide-T-cell receptor (MHCII-p-TCR) complex-mediated antigen presentation for a minimal subunit-based, multi-epitope, multistage, chemically-synthesised antimalarial vaccine is essential for inducing an appropriate immune response. Deep understanding of this MHCII-p-TCR complex's stereo-electronic characteristics is fundamental for vaccine development. This review encapsulates the main principles for achieving such epitopes' perfect fit into MHC-II human (HLADRß̞1*) or Aotus (Aona DR) molecules. The enormous relevance of several amino acids' physico-chemical characteristics is analysed in-depth, as is data regarding a 26.5 ± 2.5Å distance between the farthest atoms fitting into HLA-DRß1* structures' Pockets 1 to 9, the role of polyproline II-like (PPIIL) structures having their O and N backbone atoms orientated for establishing H-bonds with specific HLA-DRß1*-peptide binding region (PBR) residues. The importance of residues having specific charge and orientation towards the TCR for inducing appropriate immune activation, amino acids' role and that of structures interfering with PPIIL formation and other principles are demonstrated which have to be taken into account when designing immune, protection-inducing peptide structures (IMPIPS) against diseases scourging humankind, malaria being one of them.

SM-COLSARSPROT: Highly Immunogenic Supramutational Synthetic Peptides Covering the World's Population.

Fifty ~20-amino acid (aa)-long peptides were selected from functionally relevant SARS-CoV-2 S, M, and E proteins for trial B-21 and another 53 common ones, plus some new ones derived from the virus' main genetic variants for complementary trial C-21. Peptide selection was based on tremendous SARS-CoV-2 genetic variability for analysing them concerning vast human immunogenetic polymorphism for developing the first supramutational, Colombian SARS-protection (SM-COLSARSPROT), peptide mixture. Specific physicochemical rules were followed, i.e., aa predilection for polyproline type II left-handed (PPIIL) formation, replacing ß-branched, aromatic aa, short-chain backbone H-bond-forming residues, π-π interactions (n→π* and π-CH), aa interaction with π systems, and molecular fragments able to interact with them, disrupting PPIIL propensity formation. All these modified structures had PPIIL formation propensity to enable target peptide interaction with human leukocyte antigen-DRß1* (HLA-DRß1*) molecules to mediate antigen presentation and induce an appropriate immune response. Such modified peptides were designed for human use; however, they induced high antibody titres against S, M, and E parental mutant peptides and neutralising antibodies when suitably modified and chemically synthesised for immunising 61 major histocompatibility complex class II (MHCII) DNA genotyped Aotus monkeys (matched with their corresponding HLA-DRß1* molecules), predicted to cover 77.5% to 83.1% of the world's population. Such chemically synthesised peptide mixture represents an extremely pure, stable, reliable, and cheap vaccine for COVID-19 pandemic control, providing a new approach for a logical, rational, and soundly established methodology for other vaccine development.

How to Combat Gram-Negative Bacteria Using Antimicrobial Peptides: A Challenge or an Unattainable Goal?

Antimicrobial peptides (AMPs) represent a promising and effective alternative for combating pathogens, having some advantages compared to conventional antibiotics. However, AMPs must also contend with complex and specialised Gram-negative bacteria envelops. The variety of lipopolysaccharide and phospholipid composition in Gram-negative bacteria strains and species are decisive characteristics regarding their susceptibility or resistance to AMPs. Such biological and structural barriers have created delays in tuning AMPs to deal with Gram-negative bacteria. This becomes even more acute because little is known about the interaction AMP-Gram-negative bacteria and/or AMPs' physicochemical characteristics, which could lead to obtaining selective molecules against Gram-negative bacteria. As a consequence, available AMPs usually have highly associated haemolytic and/or cytotoxic activity. Only one AMP has so far been FDA approved and another two are currently in clinical trials against Gram-negative bacteria. Such a pessimistic panorama suggests that efforts should be concentrated on the search for new molecules, designs and strategies for combating infection caused by this type of microorganism. This review has therefore been aimed at describing the currently available AMPs for combating Gram-negative bacteria, exploring the characteristics of these bacteria's cell envelop hampering the development of new AMPs, and offers a perspective regarding the challenges for designing new AMPs against Gram-negative bacteria.

The First Chemically-Synthesised, Highly Immunogenic Anti-SARS-CoV-2 Peptides in DNA Genotyped Aotus Monkeys for Human Use.

Thirty-five peptides selected from functionally-relevant SARS-CoV-2 spike (S), membrane (M), and envelope (E) proteins were suitably modified for immunising MHC class II (MHCII) DNA-genotyped Aotus monkeys and matched with HLA-DRß1* molecules for use in humans. This was aimed at producing the first minimal subunit-based, chemically-synthesised, immunogenic molecules (COLSARSPROT) covering several HLA alleles. They were predicted to cover 48.25% of the world's population for 6 weeks (short-term) and 33.65% for 15 weeks (long-lasting) as they induced very high immunofluorescent antibody (IFA) and ELISA titres against S, M and E parental native peptides, SARS-CoV-2 neutralising antibodies and host cell infection. The same immunological methods that led to identifying new peptides for inclusion in the COLSARSPROT mixture were used for antigenicity studies. Peptides were analysed with serum samples from patients suffering mild or severe SARS-CoV-2 infection, thereby increasing chemically-synthesised peptides' potential coverage for the world populations up to 62.9%. These peptides' 3D structural analysis (by 1H-NMR acquired at 600 to 900 MHz) suggested structural-functional immunological association. This first multi-protein, multi-epitope, minimal subunit-based, chemically-synthesised, highly immunogenic peptide mixture highlights such chemical synthesis methodology's potential for rapidly obtaining very pure, highly reproducible, stable, cheap, easily-modifiable peptides for inducing immune protection against COVID-19, covering a substantial percentage of the human population.

Identifying HLA DRB1-DQB1 alleles associated with Chlamydia trachomatis infection and in silico prediction of potentially-related peptides.

HLA class II (HLA-II) genes' polymorphism influences the immune response to Chlamydia trachomatis (Ct), it is considered a sexually transmitted infection. However, associations between HLA-II alleles and Ct-infection have been little explored in humans; this study was thus aimed at determining HLA-DRB1-DQB1 alleles/haplotypes' effect on Ct-infection outcome in a cohort of Colombian women. Cervical sample DNA was used as template for detecting Ct by PCR and typing HLA-DRB1-DQB1 alleles/haplotypes by Illumina MiSeq sequencing. Survival models were adjusted for identifying the alleles/haplotypes' effect on Ct-outcome; bioinformatics tools were used for predicting secreted bacterial protein T- and B-cell epitopes. Sixteen HLA-DRB1 alleles having a significant effect on Ct-outcome were identified in the 262 women analysed. DRB1*08:02:01G and DRB1*12:01:01G were related to infection-promoting events. Only the DQB1*05:03:01G allele related to clearance/persistence events was found for HLA-DQB1. HLA-DRB1 allele homozygous women were associated with events having a lower probability of clearance and/or early occurrence of persistence. Twenty-seven peptides predicted in silico were associated with protective immunity against Ct; outer membrane and polymorphic membrane protein-derived peptides had regions having dual potential for being T- or B-cell epitopes. This article describes HLA-DRB1-DQB1 alleles/haplotypes related to Ct-infection resolution and the peptides predicted in silico which might probably be involved in host immune response. The data provides base information for developing future studies leading to the development of effective prevention measures against Ct-infection.

MHCBI: a pipeline for calculating peptide-MHC binding energy using semi-empirical quantum mechanical methods with explicit/implicit solvent models.

Experimentally estimating peptide-major histocompatibility complex (pMHC) binding affinity has been quite challenging due to the many receptors and the many potential ligands implicated in it. We have thus proposed a straightforward computational methodology considering the different mechanisms involved in pMHC binding to facilitate studying such receptor-ligand interactions. We have developed a pipeline using semi-empirical quantum mechanical methods for calculating pMHC class I and II molecules' binding energy (BE). This pipeline can systematize the methodology for calculating pMHC system BE, enabling the rational design of T-cell epitopes to be used as pharmaceuticals and vaccines.

The molecular basis for peptide-based antimalarial vaccine development targeting erythrocyte invasion by P. falciparum.

This work describes a methodology for developing a minimal, subunit-based, multi-epitope, multi-stage, chemically-synthesised, anti-Plasmodium falciparum malaria vaccine. Some modified high activity binding peptides (mHABPs) derived from functionally relevant P. falciparum MSP, RH5 and AMA-1 conserved amino acid regions (cHABPs) for parasite binding to and invasion of red blood cells (RBC) were selected. They were highly immunogenic as assessed by indirect immunofluorescence (IFA) and Western blot (WB) assays and protective immune response-inducers against malarial challenge in the Aotus monkey experimental model. NetMHCIIpan 4.0 was used for predicting peptide-Aotus/human major histocompatibility class II (MHCII) binding affinity in silico due to the similarity between Aotus and human immune system molecules; ∼50% of Aotus MHCII allele molecules have a counterpart in the human immune system, being Aotus-specific, whilst others enabled recognition of their human counterparts. Some peptides' 1H-NMR-assessed structural conformation was determined to explain residue modifications in mHABPs inducing secondary structure changes. These directly influenced immunological behaviour, thereby highlighting the relationship with MHCII antigen presentation. The data obtained in such functional, immunological, structural and predictive approach suggested that some of these peptides could be excellent components of a fully-protective antimalarial vaccine.

Antibodies targeting Mycobacterium tuberculosis peptides inhibit mycobacterial entry to infection target cells.

The humoral immunity regarding tuberculosis can contribute towards controlling the mycobacteria and the disease. Antigens mediating such type of immunity should thus be evaluated for formulating anti-tuberculosis vaccines. The antigen recognition of seven peptides derived from proteins on Mtb H37Rv envelope and a further seven peptides modified from them was evaluated in sera taken from people suffering Mtb infection and others free from it. Peptide sequences' ability to inhibit Mtb entry to human macrophages was determined in vitro and, after isolating peptide-specific IgG antibodies, it was ascertained which ones were exercising such inhibitory function. Aotus were inoculated with the modified peptides for evaluating the activity of the antibodies so produced. Human QTF+ and QTF- sera recognised some of the peptides and inhibited Mtb entry. The same effect was seen with peptide-specific IgG regarding all the native sequences and modified ones. Sera taken from inoculated Aotus was also able to reduce the pathogen's entry. The data showed that some peptides evaluated in this study could induce antibodies able to inhibit the pathogen's entry to human macrophages, i.e. they could represent candidates for part of an anti-tuberculosis vaccine. The methodology used here complements the evaluation of promising antigens for designing effective vaccines.

Identifying the HLA DRB1-DQB1 molecules and predicting epitopes associated with high-risk HPV infection clearance and redetection.

Several determining factors are involved in HPV infection outcomes; human leukocyte antigen (HLA) polymorphisms have been described as related factors. This study has ascertained the effect of genetic variation on HLA-DRB1 and DQB1 genes on HPV-16/-18/-31/-33/-45 and -58 clearance and redetection in Colombian women. PCR and qPCR were used for viral identification and the Illumina MiSeq system was used for HLA-typing of cervical samples (n = 276). Survival models were adjusted for identifying alleles/haplotypes related to HPV clearance/redetection; L1/L2 protein-epitope binding to MHC-II molecules was also predicted. Significant associations suggested effects favouring or hampering clearance/redetection events depending on the viral type involved in infection, e.g. just DRB1*12:01:01G favoured HPV-16 (coeff: 4.8) and HPV-45 clearance (coeff: 12.65) whilst HPV-18 (coeff: 2E-15), HPV-31 (coeff: 8E-17) and HPV-58 hindered elimination (coeff: 1E-14). An effect was only observed for some alelles when configured as haplotypes, e.g. DRB1*04:07:01G (having the greatest frequency in the target population) was associated with DQB1*02:01:1G or *03:02:03. Epitope prediction identified 23 clearance-related peptides and 29 were redetection-related; eight might have been related to HPV-16/-18 and -58 persistence and one to HPV-18 elimination. HLA allele/haplotype relationship with the course of HPV infection (clearance/redetection) depended on the infecting HPV type, in line with the specific viral epitopes displayed.

Malaria: Paving the way to developing peptide-based vaccines against invasion in infectious diseases.

Malaria remains a large-scale public health problem, killing more than 400,000 people and infecting up to 230 million worldwide, every year. Unfortunately, despite numerous efforts and research concerning vaccine development, results to date have been low and/or strain-specific. This work describes a strategy involving Plasmodium falciparum Duffy binding-like (DBL) and reticulocyte-binding protein homologue (RH) family-derived minimum functional peptides, netMHCIIpan3.2 parental and modified peptides' in silico binding prediction and modeling some Aotus major histocompatibility class II (MHCII) molecules based on known human molecules' structure to understand their differences. These are used to explain peptides' immunological behaviour when used as vaccine components in the Aotus model. Despite the great similarity between human and Aotus immune system molecules, around 50% of Aotus allele molecules lack a counterpart in the human immune system which could lead to an Aotus-specific vaccine. It was also confirmed that functional Plasmodium falciparum' conserved proteins are immunologically silent (in both the animal model and in-silico prediction); they must therefore be modified to elicit an appropriate immune response. Some peptides studied here had the desired behaviour and can thus be considered components of a fully-protective antimalarial vaccine.

p-Methoxyphenol: A potent and effective scavenger for solid-phase peptide synthesis.

During the final step of t-Boc/Bzl, solid-phase peptide synthesis (SPPS)-protecting groups from amino acids (aa) side chains must be removed from the target peptides during cleavage from the solid support. These reaction steps involve hydrolysis with hydrogen fluoride (HF) in the presence of a nucleophile (scavenger), whose function is to trap the carbocations produced during SN 1-type reactions. Five peptide sequences were synthesised for evaluating p-methoxyphenol effectiveness as a potent scavenger. After the synthesis, the resin-peptide was then separated into two equal parts to be cleaved using two scavengers: conventional reactive p-cresol (reported in the literature as an effective acyl ion eliminator) and p-methoxyphenol (hypothesised as fulfilling the same functions as the routinely used scavenger). Detailed analysis of the electrostatic potential map (EPM) revealed similarities between these two nucleophiles, regarding net atomic charge, electron density distribution, and similar pKa values. Good scavenger efficacy was observed by chromatography and mass spectrometry results for the synthesised molecules, which revealed that p-methoxyphenol can be used as a potent scavenger during SPPS by t-Boc/Bzl strategy, as similar results were obtained using the conventional scavenger.

Evaluating the immunogenicity of chemically-synthesised peptides derived from foot-and-mouth disease VP1, VP2 and VP3 proteins as vaccine candidates.

Foot-and-mouth disease (FMD) is one of the most contagious veterinary viral diseases known, having economic, social and potentially devastating environmental impacts. The vaccines currently being marketed/sold around the world for disease control and prevention in bovines do not stimulate the production of antibodies having crossed reactions to different serotypes. This means that if an animal becomes infected by a serotype which has not been included in a vaccine then it will develop the disease. Synthetic peptide vaccines represent a safer option and (depending on the design) can stimulate antibodies protecting against different variants. Based on the forgoing, this work was aimed at evaluating FMDV VP1, VP2 and VP3 protein-derived, modified and chemically-synthesised peptides' ability to induce an immune response for developing a vaccine contributing towards controlling the disease. VP1, VP2 and VP3 proteins' conserved regions were selected for this. Peptides from these regions were chemically synthesised; binding assays were then carried out for ascertaining whether they were involved in BHK-21 cell binding. Selected peptides' structure and location were studied. Peptides which did bind were modified and formulated with Montanide ISA 70 adjuvant; 17 animals were immunised twice with the formulation. The animals were genotyped by amplifying the BoLA-DRB3.2 gene. Blood samples were taken from 17 cattle on day 43 post-first immunisation for studying the formulation's immunogenicity. The sera were used in ELISA, immunofluorescence, flow cytometry, immunoadsorption and seroneutralisation assays. The A24 Cruzeiro and O1 Campos virus serotypes were used for these assays. The results revealed that even though protein exposure and 3D structure might be different amongst serotypes, the antibodies so produced could inhibit virus entry to cells, thereby showing the selected peptides' in vitro protection-inducing ability.

Assessing Peptide Binding to MHC II: An Accurate Semiempirical Quantum Mechanics Based Proposal.

Estimating peptide-major histocompatibility complex (pMHC) binding using structural computational methods has an impact on understanding overall immune function triggering adaptive immune responses in MHC class II molecules. We developed a strategy for optimizing pMHC structure interacting with water molecules and for calculating the binding energy of receptor + ligand systems, such as HLA-DR1 + HA, HLA-DR1 + CLIP, HLA-DR2 + MBP, and HLA-DR3 + CLIP, as well as a monosubstitution panel. Taking pMHC's structural properties, we assumed that ΔH ≫ -TΔS would generate a linear model for estimating relative free energy change, using three semiempirical quantum methods (PM6, PM7, and FMO-SCC-DFTB3) along with the implicit solvent models, and considering proteins in neutral and charged states. Likewise, we confirmed our approach's effectiveness in calculating binding energies having high correlation with experimental data and low root-mean-square error (<2 kcal/mol). All in all, our pipeline differentiates weak from strong peptide binders as a reliable method for studying pMHC interactions.

Designing and optimizing new antimicrobial peptides: all targets are not the same.

Because the resistance of microorganisms to the available antibiotics is a growing healthcare problem worldwide, the search for new antimicrobial peptides (AMPs) that provide useful therapeutic options has been increasing in importance. Many initial candidates have had to be discarded after having advanced to the preclinical and clinical stages. This has led to substantial losses in terms of time and money. For that reason, the essential characteristics of AMPs (i.e. their activity, selectivity, stability in physiological conditions and low production cost) must be considered in their design. In addition, peptides could be active against several kinds of cells with activity and selectivity resulting from interaction with multiple target cell components, which sometimes are present in mammalian cells as well. Thus, the cellular composition is important in the AMP-target cell interaction and must be considered in the design of AMPs, too. This review describes general aspects of AMP design, limitations concerning their therapeutic application, and optimization strategies for overcoming such limitations.

Using next-generation sequencing for characterising HLA-DRB1 and DQB1 loci in a cohort of Colombian women.

The Colombian population is characterised by a high genetic diversity, secondary to the ethnic mixture arising from colonisation. Unfortunately, few reports are available regarding HLA-DRB1 and DQB1 diversity in Colombia to date. HLA-DRB1 and DQB1 diversity was identified in this study using next-generating sequencing (NGS) on a cohort of Colombian women. Cervical samples taken from 276 women were used for typing DRB1 and DQB1 loci by Illumina MiSeq. Allele and haplotype frequencies were calculated using an expectation-maximisation algorithm. Hardy-Weinberg Equilibrium and linkage disequilibrium (LD) between loci were evaluated. Forty-seven DRB1 alleles and 14 DQB1 alleles were identified. DRB1*04:07:01G and DQB1*03:02:01G alleles occurred most frequently in the target population. Significant LD was found in 44 out of the 144 identified haplotypes, within which DRB1*04:07:01G-DQB1*03:02:01G occurred most frequently (6.56%). The alleles and haplotypes found with NGS agreed with that found in previous reports involving lower resolution for the Colombian population, and greater genetic variability was found, especially concerning DRB1. Comparing allele and haplotype frequency distribution in the target population to that of other populations denoted HLA system intra- and inter-population diversity.

Preliminary Evaluation of the Safety and Immunogenicity of an Antimalarial Vaccine Candidate Modified Peptide (IMPIPS) Mixture in a Murine Model.

Malaria continues being a high-impact disease regarding public health worldwide; the WHO report for malaria in 2018 estimated that ~219 million cases occurred in 2017, mostly caused by the parasite Plasmodium falciparum. The disease cost the lives of more than 400,000 people, mainly in Africa. In spite of great efforts aimed at developing better prevention (i.e., a highly effective vaccine), diagnosis, and treatment methods for malaria, no efficient solution to this disease has been advanced to date. The Fundación Instituto de Inmunología de Colombia (FIDIC) has been developing studies aimed at furthering the search for vaccine candidates for controlling P. falciparum malaria. However, vaccine development involves safety and immunogenicity studies regarding their formulation in animal models before proceeding to clinical studies. The present work has thus been aimed at evaluating the safety and immunogenicity of a mixture of 23 chemically synthesised, modified peptides (immune protection-inducing protein structure (IMPIPS)) derived from different P. falciparum proteins. Single and repeat dose assays were thus used with male and female BALB/c mice which were immunised with the IMPIPS mixture. It was found that single and repeat dose immunisation with the IMPIPS mixture was safe, both locally and systemically. It was observed that the antibodies so stimulated recognised the parasite's native proteins and inhibited merozoite invasion of red blood cells in vitro when evaluating the humoral immune response induced by the IMPIPS mixture. Such results suggested that the IMPIPS peptide mixture could be a safe candidate to be tested during the next stage involved in developing an antimalarial vaccine, evaluating local safety, immunogenicity, and protection in a nonhuman primate model.

The in Vitro Antigenicity of Plasmodium vivax Rhoptry Neck Protein 2 (PvRON2) B- and T-Epitopes Selected by HLA-DRB1 Binding Profile.

Malaria caused by Plasmodium vivax is a neglected disease which is responsible for the highest morbidity in both Americas and Asia. Despite continuous public health efforts to prevent malarial infection, an effective antimalarial vaccine is still urgently needed. P. vivax vaccine development involves analyzing naturally-infected patients' immune response to the specific proteins involved in red blood cell invasion. The P. vivax rhoptry neck protein 2 (PvRON2) is a highly conserved protein which is expressed in late schizont rhoptries; it interacts directly with AMA-1 and might be involved in moving-junction formation. Bioinformatics approaches were used here to select B- and T-cell epitopes. Eleven high-affinity binding peptides were selected using the NetMHCIIpan-3.0 in silico prediction tool; their in vitro binding to HLA-DRB1*0401, HLA-DRB1*0701, HLA-DRB1*1101 or HLA-DRB1*1302 was experimentally assessed. Four peptides (39152 (HLA-DRB1*04 and 11), 39047 (HLA-DRB1*07), 39154 (HLADRB1*13) and universal peptide 39153) evoked a naturally-acquired T-cell immune response in P. vivax-exposed individuals from two endemic areas in Colombia. All four peptides had an SI greater than 2 in proliferation assays; however, only peptides 39154 and 39153 had significant differences compared to the control group. Peptide 39047 was able to significantly stimulate TNF and IL-10 production while 39154 stimulated TNF production. Allele-specific peptides (but not the universal one) were able to stimulate IL-6 production; however, none induced IFN-γ production. The Bepipred 1.0 tool was used for selecting four B-cell epitopes in silico regarding humoral response. Peptide 39041 was the only one recognized by P. vivax-exposed individuals' sera and had significant differences concerning IgG subclasses; an IgG2 > IgG4 profile was observed for this peptide, agreeing with a protection-inducing role against P. falciparum and P. vivax as previously described for antigens such as RESA and MSP2. The bioinformatics results and in vitro evaluation reported here highlighted two T-cell epitopes (39047 and 39154) being recognized by memory cells and a B-cell epitope (39041) identified by P. vivax-exposed individuals' sera which could be used as potential candidates when designing a subunit-based vaccine.

Specific ß-Turns Precede PPIIL Structures Binding to Allele-Specific HLA-DRß1* PBRs in Fully-Protective Malaria Vaccine Components.

The 3D structural analysis of 62 peptides derived from highly pathogenic Plasmodium falciparum malaria parasite proteins involved in host cell invasion led to finding a striking association between particular ß-turn types located in the N-terminal peripheral flanking residue region (preceding the polyproline II left-handed structures fitting into the HLA-DRß* allele family) and modified immune protection-inducing protein structure induced long-lasting protective immunity. This is the first time association between two different secondary structures associated with a specific immunological function has been described: full, long-lasting protective immunity.