scholarly journals Identification of Continuous B-Cell Epitopes on the Protein Moiety of the 58-Kilodalton Cell Wall Mannoprotein ofCandida albicans Belonging to a Family of Immunodominant Fungal Antigens

2001 ◽  
Vol 69 (5) ◽  
pp. 2909-2919 ◽  
Author(s):  
Angel Viudes ◽  
Sofia Perea ◽  
Jose L. Lopez-Ribot
Keyword(s):  
Amino Acid ◽  
B Cell ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Amino Acid Residues ◽  
B Cell Epitopes ◽  
C Terminus ◽  
Elisa Testing ◽  
Fungal Antigens ◽  
Protein Moiety

ABSTRACT The 58-kiloDalton mannoprotein (mp58) on the surface ofCandida albicans is highly immunogenic, is expressed by allC. albicans isolates tested, and elicits strong antibody responses during candidiasis. It belongs to a family of immunodominant fungal antigens with representatives also in different species ofAspergillus. The amino acid sequence of the protein portion of mp58 as deduced from the DNA sequence of its encoding gene (FBP1/PRA1) was used to synthesize a complete set of overlapping dodecapeptides (overlap, 7; offset, 5) covalently attached to the surface of derivatized polyethylene pins. The pin-coupled peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by a number of antiserum preparations containing anti-mp58 antibodies. This comprehensive epitope-scanning study revealed the presence of multiple immunoreactive continuous B-cell epitopes within the protein sequence. Regions of increased reactivity included both the amino and carboxy termini of the mature protein (encompassing amino acid residues 16 to 50 and 286 to 299, respectively) and four internal regions spanning amino acids at positions 66 to 92, 121 to 142, 148 to 192, and 211 to 232. Further delineation of epitopic regions and identification of the boundaries of the antigenic sites was performed upon ELISA testing with a second Pepset consisting of completely overlapping 8-mer peptides spanning these reactive regions in the protein moiety of mp58. The highly reactive epitopic region at the C terminus of the protein was further evaluated using both window net and replacement net analyses. A synthetic peptide corresponding to the last 10 amino acid residues at the C terminus of the protein was immunogenic when injected into mice after being coupled to a carrier protein. Moreover, antibodies in the resulting sera specifically recognized the homologus mp58 in ELISAs and immunoblot assays. Delineation of the antibody responses to mp58 could provide the basis for the development of novel immunity-based prophylactic, therapeutic, and diagnostic techniques for the management of candidiasis.

Polymorphism of cow colostrum proteinase inhibitor

1981 ◽  
Vol 46 (3) ◽  
pp. 807-816 ◽  
Author(s):  
Věra Jonáková ◽  
Dana Čechová ◽  
Otakar Mach
Keyword(s):  
Amino Acid ◽  
Primary Structure ◽  
Proteinase Inhibitor ◽  
Protein Structures ◽  
Amino Acid Residues ◽  
Net Charge ◽  
C Terminus ◽  
Isoelectric Points ◽  
The Individual ◽  
Protein Moiety

Cow colostrum contains three isoinhibitors A, B, and C, which are glycoproteins. In this study isoinhibitor A was isolated and characterized and the structure of its protein moiety compared with the known protein structures of isoinhibitors B and C. It was found that the primary structure of isoinhibitor A is identical with the primary structure of isoinhibitor B except that the C-terminus of its molecule is shorter by five amino acid residues. Four discrete chromatographic forms (I-IV) with different isoelectric points (pI 3.8 - I, 4.0 - II, 4.3 - III, and 4.5 - IV) were isolated by chromatography on SE-Sephadex, Form I is identical with isoinhibitor A. Forms II, III, and IV are represented by mixtures of isoinhibitors A, B, and C with a heterogeneous carbohydrate moiety which affects the total net charge of the individual inhibitor forms.

scholarly journals Towards a Vaccine Against Rheumatic Fever

2006 ◽  
Vol 13 (2-4) ◽  
pp. 125-132 ◽  
Author(s):  
L. Guilherme ◽  
K. C. Faé ◽  
F. Higa ◽  
L. Chaves ◽  
S. E. Oshiro ◽  
...  
Keyword(s):  
Amino Acid ◽  
Autoimmune Disease ◽  
Rheumatic Fever ◽  
B Cell ◽  
Cell Epitope ◽  
M Protein ◽  
Bacterial Invasion ◽  
Amino Acid Residues ◽  
Terminal Portion ◽  
B Cell Epitopes

Rheumatic fever (RF) is an autoimmune disease which affects more than 20 million children in developing countries. It is triggered byStreptococcus pyogenesthroat infection in untreated susceptible individuals. Carditis, the most serious manifestation of the disease, leads to severe and permanent valvular lesions, causing chronic rheumatic heart disease (RHD). We have been studying the mechanisms leading to pathological autoimmunity in RF/RHD for the last 15 years. Our studies allowed us a better understanding of the cellular and molecular pathogenesis of RHD, paving the way for the development of a safe vaccine for a post-infection autoimmune disease. We have focused on the search for protective T and B cell epitopes by testing 620 human blood samples against overlapping peptides spanning 99 residues of the C-terminal portion of the M protein, differing by one amino acid residue. We identified T and B cell epitopes with 22 and 25 amino acid residues, respectively. Although these epitopes were from different regions of the C-terminal portion of the M protein, they showed an identical core of 16 amino acid residues. Antibodies against the B cell epitope inhibited bacterial invasion/adhesionin vitro. Our results strongly indicated that the selected T and B cell epitopes could potentially be protective againstS. pyogenes.

scholarly journals Natural immunogenic properties of bioinformatically predicted linear B-cell epitopes of dengue envelope and pre-membrane proteins

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Mahesha N. Nadugala ◽  
Chandima Jeewandara ◽  
Ramesh S. Jadi ◽  
Gathsaurie N. Malavige ◽  
Aravinda M. de Silva ◽  
...  
Keyword(s):  
Antibody Response ◽  
B Cell ◽  
Healthy Volunteers ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
B Cell Epitopes ◽  
Viral Neutralization ◽  
Denv Serotypes ◽  
Loop Region ◽  
Positive Antibody

Abstract Background The natural antibody responses to B-cell epitopes from dengue structural proteins were assessed using immune sera from people having well-defined past dengue infections with one of the four serotypes. Method Based on an immune-computational analysis previously conducted, nineteen epitopes from the envelope (E) and eight epitopes from pre-membrane (prM), which were more than 50% conserved across all the four DENV serotypes, were selected. Peptides to represent these B-cell epitopes were obtained from commercially available arrays, and were subjected to enzyme linked immunosorbent assay with sera obtained from dengue seropositive healthy volunteers (DENV1 n = 12: DENV2 n = 12: DENV3 n = 12 and DENV4 n = 12), and 10 dengue seronegative healthy volunteers from Sri Lanka. The cut-off value for the positive antibody response was set by taking the mean response of a peptide to the negative sera plus three standard deviations. The peptides (N = 7) showing the broad immune responses were used to generate antibodies in three mice (Balb/c) batches. The mice antisera were then subjected to microneutralization assays against all the four DENV serotypes. An EC50 viral neutralization ≥ 40 times the serum dilution was considered as neutralizing. Results Five of the E-peptide and two prM peptides were recognised by most individuls exposed to infections with each of the four serotypes, showing a serotype cross-reactive broad antibody response. The mice immune sera against the peptides representing the five E protein epitopes neutralized all the four DENV serotypes. Two of these five epitopes are from the Domain II, whereas one of them includes the whole bc-loop region. Conclusion The antibody responses of highly conserved epitopes across the serotypes, were broadly responsive with sera of all four DENV serotypes collected from individuals infected with only one DENV serotype. Weakly conserved epitopes showed rather specific antibody responses dominated by one or few serotypes.

scholarly journals The landscape of antibody binding in SARS-CoV-2 infection

2020 ◽  
Author(s):  
Anna S Heffron ◽  
Sean J McIlwain ◽  
Maya F Amjadi ◽  
David A Baker ◽  
Saniya Khullar ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Diagnostic Accuracy ◽  
B Cell ◽  
Immunosorbent Assay ◽  
Antibody Binding ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Peptide Microarray ◽  
B Cell Epitopes ◽  
Homologous Peptide

The search for potential antibody-based diagnostics, vaccines, and therapeutics for pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has focused almost exclusively on the spike (S) and nucleocapsid (N) proteins. Coronavirus membrane (M), ORF3a, and ORF8 proteins are humoral immunogens in other coronaviruses (CoVs) but remain largely uninvestigated for SARS-CoV-2. Here we use ultradense peptide microarray mapping to show that SARS-CoV-2 infection induces robust antibody responses to epitopes throughout the SARS-CoV-2 proteome, particularly in M, in which one epitope achieved excellent diagnostic accuracy. We map 79 B cell epitopes throughout the SARS-CoV-2 proteome and demonstrate that antibodies that develop in response to SARS-CoV-2 infection bind homologous peptide sequences in the six other known human CoVs. We also confirm reactivity against four of our top-ranking epitopes by enzyme-linked immunosorbent assay (ELISA). Illness severity correlated with increased reactivity to nine SARS-CoV-2 epitopes in S, M, N, and ORF3a in our population. Our results demonstrate previously unknown, highly reactive B cell epitopes throughout the full proteome of SARS-CoV-2 and other CoV proteins.

scholarly journals The landscape of antibody binding in SARS-CoV-2 infection

PLoS Biology ◽  
2021 ◽  
Vol 19 (6) ◽  
pp. e3001265
Author(s):  
Anna S. Heffron ◽  
Sean J. McIlwain ◽  
Maya F. Amjadi ◽  
David A. Baker ◽  
Saniya Khullar ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Diagnostic Accuracy ◽  
B Cell ◽  
Immunosorbent Assay ◽  
Antibody Binding ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Peptide Microarray ◽  
B Cell Epitopes ◽  
Homologous Peptide

The search for potential antibody-based diagnostics, vaccines, and therapeutics for pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has focused almost exclusively on the spike (S) and nucleocapsid (N) proteins. Coronavirus membrane (M), ORF3a, and ORF8 proteins are humoral immunogens in other coronaviruses (CoVs) but remain largely uninvestigated for SARS-CoV-2. Here, we use ultradense peptide microarray mapping to show that SARS-CoV-2 infection induces robust antibody responses to epitopes throughout the SARS-CoV-2 proteome, particularly in M, in which 1 epitope achieved excellent diagnostic accuracy. We map 79 B cell epitopes throughout the SARS-CoV-2 proteome and demonstrate that antibodies that develop in response to SARS-CoV-2 infection bind homologous peptide sequences in the 6 other known human CoVs. We also confirm reactivity against 4 of our top-ranking epitopes by enzyme-linked immunosorbent assay (ELISA). Illness severity correlated with increased reactivity to 9 SARS-CoV-2 epitopes in S, M, N, and ORF3a in our population. Our results demonstrate previously unknown, highly reactive B cell epitopes throughout the full proteome of SARS-CoV-2 and other CoV proteins.

scholarly journals Primary- and secondary-structural analysis of a unique prokaryotic metallothionein from a Synechococcus sp. cyanobacterium

1988 ◽  
Vol 251 (3) ◽  
pp. 691-699 ◽  
Author(s):  
R W Olafson ◽  
W D McCubbin ◽  
C M Kay
Keyword(s):  
Amino Acid ◽  
Metal Binding ◽  
Helical Structure ◽  
Basic Amino Acid ◽  
Cysteine Residues ◽  
Amino Acid Residues ◽  
C Terminus ◽  
Empirical Relations ◽  
Heavy Metal Binding ◽  
Synechococcus Sp

Biochemical and physiological studies of Synechococcus cyanobacteria have indicated the presence of a low-Mr heavy-metal-binding protein with marked similarity to eukaryotic metallothioneins (MTs). We report here the characterization of a Synechococcus prokaryotic MT isolated by gel-permeation and reverse-phase chromatography. The large number of variants of this molecule found during chromatographic separation could not be attributed to the presence of major isoproteins as assessed by amino acid analysis and amino acid sequencing of isoforms. Two of the latter were shown to have identical primary structures that differed substantially from the well-described eukaryotic MTs. In addition to six long-chain aliphatic residues, two aromatic residues were found adjacent to one another near the centre of the molecule, making this the most hydrophobic MT to be described. Other unusual features included a pair of histidine residues located in repeating Gly-His-Thr-Gly sequences near the C-terminus and a complete lack of association of hydroxylated residues with cysteine residues, as is commonly found in eukaryotes. Similarly, aside from a single lysine residue, no basic amino acid residues were found adjacent to cysteine residues in the sequence. Most importantly, sequence alignment analyses with mammalian, invertebrate and fungal MT sequences showed no statistically significant homology aside from the presence of Cys-Xaa-Cys structures common to all MTs. On the other hand, like other MTs, the prokaryotic molecule appears to be free of alpha-helical structure but has a considerable amount of beta-structure, as predicted by both c.d. measurements and the Chou & Fasman empirical relations. Considered together, these data suggested that some similarity between the metal-thiolate clusters of the prokaryote and eukaryote MTs may exist.

scholarly journals Deletion of the Actin-Associated Tropomyosin Tpm3 Leads to Reduced Cell Complexity in Cultured Hippocampal Neurons—New Insights into the Role of the C-Terminal Region of Tpm3.1

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 715
Author(s):  
Tamara Tomanić ◽  
Claire Martin ◽  
Holly Stefen ◽  
Esmeralda Parić ◽  
Peter Gunning ◽  
...  
Keyword(s):  
Amino Acid ◽  
Hippocampal Neurons ◽  
Molecular Mechanisms ◽  
Growth Cones ◽  
Amino Acid Residues ◽  
Terminal Amino Acid ◽  
C Terminus ◽  
Primary Mouse ◽  
Terminal Amino

Tropomyosins (Tpms) have been described as master regulators of actin, with Tpm3 products shown to be involved in early developmental processes, and the Tpm3 isoform Tpm3.1 controlling changes in the size of neuronal growth cones and neurite growth. Here, we used primary mouse hippocampal neurons of C57/Bl6 wild type and Bl6Tpm3flox transgenic mice to carry out morphometric analyses in response to the absence of Tpm3 products, as well as to investigate the effect of C-terminal truncation on the ability of Tpm3.1 to modulate neuronal morphogenesis. We found that the knock-out of Tpm3 leads to decreased neurite length and complexity, and that the deletion of two amino acid residues at the C-terminus of Tpm3.1 leads to more detrimental changes in neurite morphology than the deletion of six amino acid residues. We also found that Tpm3.1 that lacks the 6 C-terminal amino acid residues does not associate with stress fibres, does not segregate to the tips of neurites, and does not impact the amount of the filamentous actin pool at the axonal growth cones, as opposed to Tpm3.1, which lacks the two C-terminal amino acid residues. Our study provides further insight into the role of both Tpm3 products and the C-terminus of Tpm3.1, and it forms the basis for future studies that aim to identify the molecular mechanisms underlying Tpm3.1 targeting to different subcellular compartments.

scholarly journals Multiple Distinct Sets of Stereotyped Antigen Receptors Indicate a Role for Antigen in Promoting Chronic Lymphocytic Leukemia

2004 ◽  
Vol 200 (4) ◽  
pp. 519-525 ◽  
Author(s):  
Bradley T. Messmer ◽  
Emilia Albesiano ◽  
Dimitar G. Efremov ◽  
Fabio Ghiotto ◽  
Steven L. Allen ◽  
...  
Keyword(s):  
Amino Acid ◽  
B Cell ◽  
Structural Features ◽  
Lymphocytic Leukemia ◽  
Cell Subpopulation ◽  
Amino Acid Residues ◽  
Antigen Receptors ◽  
Region Gene ◽  
V Region ◽  
Unique Amino Acid

Previous studies suggest that the diversity of the expressed variable (V) region repertoire of the immunoglobulin (Ig)H chain of B-CLL cells is restricted. Although limited examples of marked constraint in the primary structure of the H and L chain V regions exist, the possibility that this level of restriction is a general principle in this disease has not been accepted. This report describes five sets of patients, mostly with unmutated or minimally mutated IgV genes, with strikingly similar B cell antigen receptors (BCRs) arising from the use of common H and L chain V region gene segments that share CDR3 structural features such as length, amino acid composition, and unique amino acid residues at recombination junctions. Thus, a much more striking degree of structural restriction of the entire BCR and a much higher frequency of receptor sharing exists among patients than appreciated previously. The data imply that either a significant fraction of B-CLL cells was selected by a limited set of antigenic epitopes at some point in their development and/or that they derive from a distinct B cell subpopulation with limited Ig V region diversity. These shared, stereotyped Ig molecules may be valuable probes for antigen identification and important targets for cross-reactive idiotypic therapy.

scholarly journals Investigation of major amino acid residues of anti-norfloxacin monoclonal antibodies responsible for binding with fluoroquinolones

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Patamalai Boonserm ◽  
Songchan Puthong ◽  
Thanaporn Wichai ◽  
Sajee Noitang ◽  
Pongsak Khunrae ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Amino Acid ◽  
3D Structure ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Amino Acid Residues ◽  
Variable Regions ◽  
Complementarity Determining Regions ◽  
Crucial Part ◽  
Major Amino Acid

AbstractIt is important to understand the amino acid residues that govern the properties of the binding between antibodies and ligands. We studied the binding of two anti-norfloxacins, anti-nor 132 and anti-nor 155, and the fluoroquinolones norfloxacin, enrofloxacin, ciprofloxacin, and ofloxacin. Binding cross-reactivities tested by an indirect competitive enzyme-linked immunosorbent assay indicated that anti-nor 132 (22–100%) had a broader range of cross-reactivity than anti-nor 155 (62–100%). These cross-reactivities correlated with variations in the numbers of interacting amino acid residues and their positions. Molecular docking was employed to investigate the molecular interactions between the fluoroquinolones and the monoclonal antibodies. Homology models of the heavy chain and light chain variable regions of each mAb 3D structure were docked with the fluoroquinolones targeting the crucial part of the complementarity-determining regions. The fluoroquinolone binding site of anti-nor 155 was a region of the HCDR3 and LCDR3 loops in which hydrogen bonds were formed with TYR (H:35), ASN (H:101), LYS (H:106), ASN (L:92), and ASN (L:93). These regions were further away in anti-nor 132 and could not contact the fluoroquinolones. Another binding region consisting of HIS (L:38) and ASP (H:100) was found for norfloxacin, enrofloxacin, and ciprofloxacin, whereas only ASP (H:100) was found for ofloxacin.

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