Molecular analysis of peptides from the GH loop of foot-and-mouth disease virus C-S30 using surface plasmon resonance: a role for kinetic rate constants

A foot-and-mouth disease virus (FMDV) field variant, isolate C-S30 (also named C-1-Barcelona), is known to contain four changes within the main antigenic site A (GH loop of capsid protein VP1, residues 136-150), at least one of which (Leu147 --> Val) involves a highly conserved position, critical for antibody recognition in the reference strain C-S8c1. However, immunoenzymatic analysis of FMDV C-S30 showed it was recognised by 4C4, a monoclonal antibody that specifically targets site A. This remarkable behaviour has led us to analyse the individual and combined contributions of the four mutations to the antigenicity of C-S30, by surface plasmon resonance (SPR) and enzyme-linked immunosorbent assay (ELISA) studies of pentadecapeptides displaying all possible combinations of the four replacements. Analysis of this family of C-S30-derived analogues shows a certain level of antibody recognition by SPR. In addition, SPR data suggest that kinetic rate constants provide an indirect measure, on the one hand, of paratope accessibility (association rate constant) and, on the other hand, of peptide fitness to the same paratope (dissociation rate constant)

Surface plasmon resonance screening of synthetic peptides mimicking the immunodominant region of C-S8c1 foot-and-mouth disease virus

The main antigenic site (site A) of foot-and-mouth disease virus (FMDV, strain C-S8c1) may be adequately reproduced by a 15-peptide with the amino acid sequence H-YTASARGDLAHLTTT-NH2 (A15), corresponding to the residues 136-150 of the viral protein VPI. The effect of amino acid substitutions within A15 on its antigenicity towards monoclonal antibodies (MAb) raised against antigenic site A, has been studied by means of BIAcore technology, based on surface plasmon resonance (SPR). Although these antigenicities have previously been determined from enzyme-linked immunosorbent assays (ELISA), the SPR-based technique is superior in that it allows a fast and straightforward screening of antigens while simultaneously providing kinetic data of the antigen-antibody interaction. With a view to screening fairly large libraries of individual peptides, we have inverted the typical SPR experiment by immobilizing the MAb on the sensor surface and using peptides as soluble analytes. We report the validation of this approach through the screening of 44 site A peptides, with results generally in good agreement with the relative antigenicities previously determined by competition ELISA

Antigenicity modulation upon peptide cyclization: application to the GH loop of foot-and-mouth disease virus strain C-1-Barcelona

Foot-and-mouth disease virus (FMDV) isolate C-1-Barcelona (or C-S30) includes four replacements within its immunodominant site (GH loop, residues 136-150 of capsid protein VP1, YTTSTRGDLAHVTAT), relative to reference strain C-S8cl (YTASAR-GDLAHLTTT). Although one of the mutations in C-S30 ((147)Leu --> Val) is known to be detrimental for antibody recognition, reactivity of this isolate with the neutralizing monoclonal antibody (mAb) 4C4, raised against FMDV C-1-Brescia (GH loop: YTASTRGDLAHLTAT), was indistinguishable from those of strains C-S8cl or C-1-Brescia. A structural interpretation for these somewhat striking findings is available, based on the observation that 15-residue peptides reproducing the C-S30 and C-S8cl GH loops adopt very similar, quasi-circular, conformations in crystal complexes with 4C4. Nevertheless, surface plasmon resonance (SPR) kinetic analyses of the interactions between these peptides and three anti-GH loop mAbs have now revealed that the linear C-S30 peptides were less antigenic in solution than their C-S8cl and C-1-Brescia counterparts. We have, therefore, tried to modulate peptide antigenicity in solution by cyclization. Functional SPR and structural two dimensional proton nuclear magnetic resonance (2D-H-1 NMR) studies of both linear and cyclic peptide antigens are discussed here. Conformation seems to have an important role in peptide antigenicity, even when continuous (i.e. linear) antigenic sites are involved

Direct single-step surface plasmon resonance analysis of interactions between small peptides and immobilized monoclonal antibodies

Surface plasmon resonance (SPR) methods have been optimized to permit direct kinetic analysis of the antigenic peptide analytes interacting with immobilized monoclonal antibodies (mAbs). High reproducibility and a significant correlation between SPR and previous ELISA data on the same set of antibodies and peptides were observed. The kinetic data obtained provide further insight into the structure of the main antigenic site of foot-and-mouth disease virus (FMDV)

Fibroblast growth factor 21 in breast milk controls neonatal intestine function

FGF21 is a hormonal factor with important functions in the control of metabolism. FGF21 is found in rodent and human milk. Radiolabeled FGF21 administered to lactating dams accumulates in milk and is transferred to neonatal gut. The small intestine of neonatal (but not adult) mice highly expresses β-Klotho in the luminal area. FGF21-KO pups fed by FGF21-KO dams showed decreased expression and circulating levels of incretins (GIP and GLP-1), reduced gene expression of intestinal lactase and maltase-glucoamylase, and low levels of galactose in plasma, all associated with a mild decrease in body weight. When FGF21-KO pups were nursed by wild-type dams (expressing FGF21 in milk), intestinal peptides and digestive enzymes were up-regulated, lactase enzymatic activity was induced, and galactose levels and body weight were normalized. Neonatal intestine explants were sensitive to FGF21, as evidenced by enhanced ERK1/2 phosphorylation. Oral infusion of FGF21 into neonatal pups induced expression of intestinal hormone factors and digestive enzymes, lactase activity and lactose absorption. These findings reveal a novel role of FGF21 as a hormonal factor contributing to neonatal intestinal function via its presence in maternal milk. Appropriate signaling of FGF21 to neonate is necessary to ensure optimal digestive and endocrine function in developing intestine

Epimacular brachytherapy for wet AMD: current perspectives

Age-related macular degeneration (AMD) is considered the most common cause of blindness in the over-60 age group in developed countries. There are basically two forms of presentation: geographic (dry or atrophic) and wet (neovascular or exudative). Geographic atrophy accounts for approximately 85%-90% of ophthalmic frames and leads to a progressive degeneration of the retinal pigment epithelium and the photoreceptors. Wet AMD causes the highest percentage of central vision loss secondary to disease. This neovascular form involves an angiogenic process in which newly formed choroidal vessels invade the macular area. Today, intravitreal anti-angiogenic drugs attempt to block the angiogenic events and represent a major advance in the treatment of wet AMD. Currently, combination therapy for wet AMD includes different forms of radiation delivery. Epimacular brachytherapy (EMBT) seems to be a useful approach to be associated with current anti-vascular endothelial growth factor agents, presenting an acceptable efficacy and safety profile. However, at the present stage of research, the results of the clinical trials carried out to date are insufficient to justify extending routine use of EMBT for the treatment of wet AMD