Biomedical subjects

Fanny Guzmán

Publications and source records attributed to Fanny Guzmán.

13 recordsLinked to original sources

PubMed2006

BTM-P1 polycationic peptide biological activity and 3D-dimensional structure.

The novel BTM-P1 peptide interferes with energetic processes in mitochondria; its antimicrobial activity against Gram-positive and Gram-negative bacteria is described here. BTM-P1 three-dimensional structure was determined by 1H NMR to explain its biological mechanisms and membrane activity. Structural data indicated that BTM-P1 can form an alpha-helix; circular dichroism analysis confirmed the peptide's propensity to behave as a typical transmembrane helix in a lipidic environment. According to the structural characteristics of the polycationic BTM-P1 peptide so revealed, its biological activity can be explained by a mechanism involving the formation of ion-permeable channels in biomembranes.

Amino Acid Sequence↗

PubMed2006

Immune response in mice and cattle after immunization with a Boophilus microplus DNA vaccine containing bm86 gene.

Plasmid pBMC2 encoding antigen Bm86 from a Colombian strain of cattle tick Boophilus microplus, was used for DNA-mediated immunization of BALB/c mice, employing doses of 10 and 50microg, delivered by intradermic and intramuscular routes. Anti-Bm86 antibody levels were significantly higher compared to control mice treated with PBS. In the evaluation of immunoglobulin isotypes, significant levels of IgG2a and IgG2b were observed in mice immunized with 50microg of pBMC2. Measurement of interleukine (IL) levels (IL-4, IL-5, IL-12(p40)) and interferon-gamma (IFN-gamma) in the sera of mice immunized with pBMC2 indicated high levels of IL-4 and IL-5, although there were also significant levels of IFN-gamma. Mice immunized with pBMC2 showed antigen-specific stimulation of splenocytes according to the incorporation of bromodeoxyuridine and IFN-gamma secretion. In all trials, mice injected intramuscularly with 50microg of pBMC2 presented the highest immune response. Moreover, cattle immunized with this DNA vaccine showed antibody production significantly different to the negative control. In conclusion, these results suggest the potential of DNA immunization with pBMC2 to induce humoral and cellular immune responses against B. microplus.

Animals↗

PubMed2004

Specific erythrocyte binding capacity and biological activity of Plasmodium falciparum-derived rhoptry-associated protein 1 peptides.

Rhoptry-associated protein 1 (RAP1) is a merozoite antigen within Plasmodium falciparum rhoptries as yet having no specific function described for it. Synthetic peptides spanning the RAP1 sequence were tested in erythrocyte binding assays to identify possible RAP1 functional regions. Five high activity binding peptides (HABPs) were identified; 26201, 26202, 26203 and 26204 spanned residues 461C-K540 within RAP1 Cys region, whilst 26188 (201T-Y220) was located in p67 amino terminal. The results showed that peptide binding was saturable, some HABPs inhibited in vitro merozoite invasion and specifically bound to a 72 kDa protein in red blood cell membrane. HABP possible function in merozoite invasion of erythrocytes is also discussed.

Amino Acid Sequence↗

PubMed2004

Mapping the anatomy of a Plasmodium falciparum MSP-1 epitope using pseudopeptide-induced mono- and polyclonal antibodies and CD and NMR conformation analysis.

Antigen structure modulation represents an approach towards designing subunit malaria vaccines. A specific epitope's alpha carbon stereochemistry, as well as its backbone topochemistry, was assessed for obtaining novel malarial immunogens. A variety of MSP-1(38-61) Plasmodium falciparum epitope pseudopeptides derived were synthesised, based on solid-phase pseudopeptide chemistry strategies; these included all-L, all-D, partially-D substituted, all-Psi-[NH-CO]-Retro, all-Psi-[NH-CO]-Retro-inverso, and Psi-[CH2NH] reduced amide surrogates. We demonstrate that specific recombinant MSP-1(34-469) fragment binding to red blood cells (RBCs) is specifically inhibited by non-modified MSP-1(42-61), as well as by its V52-L53, M51-V52 reduced amide surrogates and partial-D substitutions in K48 and E49. In vivo tests revealed that reduced amide pseudopeptide-immunised Aotus monkeys induced neutralising antibodies specifically recognising the MSP-1 N-terminus region. These findings support the role of molecular conformation in malaria vaccine development.

Amino Acid Sequence↗

PubMed2003

Human papillomavirus type 16 and 18 L1 protein peptide binding to VERO and HeLa cells inhibits their VLPs binding.

Human papillomaviruses (HPVs) are the cause of epithelial lesions, HPV type 16 and type 18 being associated with the development of anogenital cancer. The L1 Major Capsid Protein (L1) represents about 90% of total HPV protein and is involved in virus-host cell interaction, but little is known about this binding process. L1 sequences from HPV types 16 and 18 were synthesized in 56 20-mer peptides, covering the entire protein, HPLC-purified, (125)I-radiolabeled and tested in VERO and HeLa cell-binding assays to identify those peptides with high specific binding activity. Peptides 18283 (residues 54-77) and 18294 (274-308) from HPV16 L1, as well as 18312 (59-78) and 18322 (259-278) from HPV18 L1, presented high specific target cell binding activity. Peptide 18283 and 18294 affinity constants were 300 and 600 nM, respectively. Enzyme cell treatment before binding assay indicated that VERO and HeLa cell peptide receptor is a surface-exposed protein. There was a 60% reduction in peptide 18283 binding to heparin lyase-treated cells. Cross-linking assays showed that these proteins molecular weights were around 69 and 54 kDa. Peptides 18283 and 18294 specifically inhibited HPV-16 VLP binding to HeLa cells. According to the L1- and VLP-reported structure, both peptides are close on the VLP-surface, belonging to the outer surface broad pockets suggested as being potential receptor sites. Furthermore, it has been reported that a conserved motif from peptide 18294 is the target for neutralizing antibodies. These results suggest that such binding sequences are used by the virus as cell-binding regions.

Amino Acid Sequence↗

PubMed2003

Immunogenicity of Sm32 synthetic peptides derived from the Schistosoma mansoni adult worm.

The previously called "hemoglobinase" Sm32 molecule of the adult worm of Schistosoma mansoni was chemically synthesized in 22 polymeric peptides based on the t-boc strategy. Their immunogenicity was evaluated in rabbits to which a mixture of five to six peptides of 20 amino acids long were given in three doses with Freund's adjuvant. Seventeen peptides were found to be immunogenic, and sera from immunized rabbits corresponding to the molecule from the first 335 amino acids, recognized the 32 kDa native protein from the adult worm antigen by western blot. Of those, the relevant peptides responsible of the recognition of the original molecule corresponded to amino acids 101-120, 121-140 and 244-268, based on inhibition competitive assays. Because Sm32 is one of the excretory and secretory molecules released with the vomitus of the adult worm, it is one of the target antigens for detection in plasma of infected individuals. The production of these polyclonal monospecific antibodies against the synthetic peptides could be of value in the immunodiagnosis of this parasitosis.

Aging↗

PubMed2003

MHC allele-specific binding of a malaria peptide makes it become promiscuous on fitting a glycine residue into pocket 6.

Peptide 1585 (EVLYLKPLAGVYRSLKKQLE) has a highly conserved amino-acid sequence located in the Plasmodium falciparum main merozoite surface protein (MSP-1) C-terminal region, required for merozoite entry into human erythrocytes and therefore represents a vaccine candidate for P. falciparum malaria. Original sequence-specific binding to five HLA DRB1* alleles (0101, 0102, 0401, 0701, and 1101) revealed this peptide's specific HLA DRB1*0102 allele binding. This peptide's allele-specific binding to HLA DRB1*0102 took on broader specificity for the DRB1*0101, -0401, and -1101 alleles when lysine was replaced by glycine in position 17 (peptide 5198: EVLYLKPLAGVYRSLKG(17)QLE). Binding of the identified G(10)VYRSLKGQLE(20) C-terminal register to these alleles suggests that peptide promiscuous binding relied on fitting Y(12), L(15), and G(17) into P-1, P-4, and P-6, respectively. The implications of the findings and the future of this synthetic vaccine candidate are discussed.

Alleles↗

PubMed2003

Expression, polymorphism analysis, reticulocyte binding and serological reactivity of two Plasmodium vivax MSP-1 protein recombinant fragments.

Among the four parasite species causing malaria in humans, Plasmodium vivax prevails on both the Asian and the American continents. Several antigens from this parasite's erythrocytic stages have been characterised and some of them are considered to be good vaccine candidates. The P. vivax merozoite surface protein-1 (PvMSP-1) is a 200 kDa antigen, thought to mediate the initial contact between the merozoite and the erythrocyte. An effective blockage of this interaction could be important in anti-malarial vaccine design. This study analyses the genetic polymorphism, binding to both reticulocytes and erythrocytes, antigenicity and immunogenicity of two recombinant proteins belonging to the 33 kDa PvMSP-1 proteolytic fragment. Both regions showed very low genetic variation, bound reticulocytes with higher affinity than erythrocytes, were recognised by naturally P. vivax-infected patient sera and were immunogenic when used to immunise rabbits, making them good vaccine candidates against P. vivax, to be further preclinically tested in the Aotus monkey model.

Amino Acid Sequence↗

PubMed2003

Plasmodium falciparum normocyte binding protein (PfNBP-1) peptides bind specifically to human erythrocytes.

Plasmodium falciparum normocyte binding protein-1 (PfNBP-1), a Plasmodium vivax RBP-1 orthologue is expressed in the apical merozoite area. PfNBP-1 binds directly to human erythrocyte membrane in a sialic acid-dependent but trypsin-resistant way. Erythrocyte binding assays were done with synthetic peptides covering the sequence reported as PfNBP-1. Two specific erythrocyte high activity binding peptides were found: 101VFINDLDTYQYEYFYEWNQ(120), peptide 26332, and 181NTKETYLKELNKKKMLQNKK(200), peptide 26336. These two peptides' binding was saturable and presenting nanomolar affinity constants. The critical binding residues (those residues underlined and highlighted in bold) were determined by competition assays with glycine-scan analogue peptides. These peptides were able to block merozoite in vitro invasion of erythrocytes.

Amino Acid Sequence↗

PubMed2003

Analysis of a Plasmodium falciparum EBA-175 peptide with high binding capacity to erythrocytes and their analogues using 1H NMR.

A 175-erythrocyte-binding protein (EBA-175) conserved high-activity binding peptide (HABP), called 1783 (nonimmunogenic, nonprotective against Plasmodium falciparum malaria), was analyzed for antigenic and protective activity in Aotus monkeys, together with several of its analogues. 1H NMR studies of peptides 17912, 14016, and 22814 allowed their structure to be related to their biological function. These peptides showed helical regions having differences in their position and length. Nonimmunogenic, nonprotective peptides 1783 and 17912 showed an extensive helical region, while the 22814 immunogenic protective peptide's alpha-helix was found in the N-terminal region. This suggests that the more flexible C-terminal region will allow better interaction between these peptides and immune system molecules as well as relating these peptides' three-dimensional structure to their immunogenicity and protective activity, thus leading to a more rational development of the new malaria multicomponent vaccine.

Amino Acid Sequence↗

PubMed2003

Characterizing cellular immune response to kinetoplastid membrane protein-11 (KMP-11) during Leishmania (Viannia) panamensis infection using dendritic cells (DCs) as antigen presenting cells (APCs).

In vitro peptide binding assays and DCs pulsed with recombinant KMP-11 (rKMP-11) plus six 20-mer overlapping peptides covering the entire protein of Leishmania (Viannia) panamensis (L(V)p) promastigotes were used to identify T-cell epitopes in this protein. Such in vitro binding assays, using HLA DRB1* 0101, -0401, -0701 and -1101 alleles, demonstrated that two peptide sequences (DEEFNKKMQEQNAKFFADKP and FKHKFAELLEQQKAAQYPSK) exhibited high HLA DRB1* 0401 allele binding capacity. rKMP-11 specific T-cell proliferation and cytokine production, derived from 13 volunteers exposed to the parasite, suggested that using autologous DCs as APCs becomes advantageous in uncovering T-cell epitopes promoting proliferation and differences in IFN-gamma and IL-4 production in T-cells from volunteers with ACTIVE and CURED undetectable disease when other APCs were used. The two peptides which bound in vitro to the HLA DRB1* 0401 allele were immunogenic in HLA DRB1* 04 volunteers, thus validating the use of in vitro binding assays for predicting epitopes in this protein. The experimental approach used here may prove useful for characterizing T-cell epitopes in a protein useful in designing peptide-based vaccine candidates for Leishmania and other intracellular pathogens.

Amino Acid Sequence↗

PubMed2002

Plasmodium vivax promiscuous T-helper epitopes defined and evaluated as linear peptide chimera immunogens.

Clinical trials of malaria vaccines have confirmed that parasite-derived T-cell epitopes are required to elicit consistent and long-lasting immune responses. We report here the identification and functional characterization of six T-cell epitopes that are present in the merozoite surface protein-1 of Plasmodium vivax (PvMSP-1) and bind promiscuously to four different HLA-DRB1* alleles. Each of these peptides induced lymphoproliferative responses in cells from individuals with previous P. vivax infections. Furthermore, linear-peptide chimeras containing the promiscuous PvMSP-1 T-cell epitopes, synthesized in tandem with the Plasmodium falciparum immunodominant circumsporozoite protein (CSP) B-cell epitope, induced high specific antibody titers, cytokine production, long-lasting immune responses, and immunoglobulin G isotype class switching in BALB/c mice. A linear-peptide chimera containing an allele-restricted P. falciparum T-cell epitope with the CSP B-cell epitope was not effective. Two out of the six promiscuous T-cell epitopes exhibiting the highest anti-peptide response also contain B-cell epitopes. Antisera generated against these B-cell epitopes recognize P. vivax merozoites in immunofluorescence assays. Importantly, the anti-peptide antibodies generated to the CSP B-cell epitope inhibited the invasion of P. falciparum sporozoites into human hepatocytes. These data and the simplicity of design of the chimeric constructs highlight the potential of multimeric, multistage, and multispecies linear-peptide chimeras containing parasite promiscuous T-cell epitopes for malaria vaccine development.

Amino Acid Sequence↗

PubMedDate not supplied

Immunogenicity and protectivity of Plasmodium falciparum EBA-175 peptide and its analog is associated with alpha-helical region shortening and displacement.

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.

Amino Acid Sequence↗