scholarly journals Identification of antigenic epitopes for Guertu virus nucleocapsid protein

2021 ◽  
Author(s):  
Siyuan Wang ◽  
Xihong Yue ◽  
Alai Shalitanati ◽  
Abulimiti Moming ◽  
Shu Shen ◽  
...  
Keyword(s):  
Amino Acid ◽  
Epitope Mapping ◽  
Virus Detection ◽  
Polyclonal Antiserum ◽  
Detection Methods ◽  
Amino Acid Residues ◽  
High Conservation ◽  
Potential Pathogenicity ◽  
Severe Fever ◽  
Antigenic Epitopes

Abstract Guertu virus (GTV), a novel tick-borne virus with potential pathogenicity, was first isolated from Dermacentor nuttalli in Xinjiang, China, in 2014. GTV has been shown to infect animal and human cell lines and to be pathogenic in mice. The viral nucleoprotein (NP) is the most conserved immunogenic protein. Elucidating the B-cell epitopes (BCEs) in the immunodominant region of the NP is important for the development of virus detection methods and vaccines. In order to identify the minimal motifs of linear BCEs in the NP of the GTV DXM strain, we used an improved biosynthetic peptide (BSP) method to truncate GTV NP into 30 16mer-peptides with 8 overlapping amino acid residues spanning the full length of the protein. The peptides were analyzed by western blot using rabbit anti-GTV NP polyclonal antiserum, and four positive 16mer-peptides were obtained. The 16mer-peptides were then truncated into 31 8mer-peptides with 7 overlapping amino acid residues and 10mer-peptides with 9 overlapping amino acid residues to screen for BCEs that can react with the rabbit anti-GTV NP polyclonal antiserum. The results showed that there were 6 minimal BCE motifs, namely, Enp1, 88EKYGLVER95; Enp2, 88EKYGLVER95; Enp3, 162TTKILMEA169; Enp4, 187GASKAEVY194; Enp5, 191AEVYNSFR198; and Enp6, 236ETAAAAYRNL245. Positive sheep sera could recognize all six BCEs with anti-GTV antibodies. The BCEs were aligned with the sequences of eight representative severe fever with thrombocytopenia syndrome phlebovirus strains from different countries and regions that were evolutionarily closely related to GTV. The sequence identity of the BCEs ranged from 80–100%, thus showing high conservation. The fine epitope mapping of GTV NP can be used to explore the biological and immunological properties of GTV NP antigens and serve as basic data for the development of multi-epitope detection reagents and vaccine design for GTV.

scholarly journals Mapping of Escherichia coli H27-Specific Epitope from H-Specific Polypeptides

2001 ◽  
Vol 8 (6) ◽  
pp. 1126-1130
Author(s):  
J. N. Seah ◽  
J. Kwang
Keyword(s):  
Amino Acid ◽  
Polyclonal Antibodies ◽  
Variable Region ◽  
Antigenic Determinants ◽  
Amino Acid Residues ◽  
Flagellin Gene ◽  
Specific Epitope ◽  
Central Variable Region ◽  
Mab Production ◽  
Antigenic Epitopes

ABSTRACT A murine monoclonal antibody (MAb) reactive to H27 flagellin antigen was produced and characterized. Forty-nine partially purified native H-type flagellins were used to evaluate the specificity of the MAb. The fliC gene of H27 is 1,464 bp in length (487 amino acids [aa]; 50.88 kDa). The central variable region (CVR) of the H27 flagellin gene was defined by comparison with flagellin sequences derived from H8, H34, and H49. To study the distribution of antigenic epitopes, the CVR covering amino acid residues 70 to 457 (388 aa) was dissected into seven overlapping fragments. Fragments carrying the H-type-specific antigenic determinants were identified by H27-specific antiserum. Polyclonal antibodies raised against different H-type flagellin proteins were used to determine the cross-reactive determinants. Three fragments, spanning amino acid residues 240 to 380, which carried the potential H-specific determinants were used for MAb production. A MAb specific to H27 was produced, and the specific epitope was mapped to amino acid residues 330 to 340. In this study, we produced MAbs from predetermined H27-specific polypeptides and used whole flagellin in enzyme-linked immunosorbent assays to circumvent the interference of anti-glutathione S-transferase antibodies. These factors when combined could help to improve the identification of the desired MAb.

The human anion transport protein, band 3, contains a CD36-like binding domain forPlasmodium falciparum-infected erythrocytes

Parasitology ◽  
1996 ◽  
Vol 112 (3) ◽  
pp. 261-267 ◽  
Author(s):  
I. Crandall ◽  
I. W. Sherman
Keyword(s):  
Monoclonal Antibody ◽  
Amino Acid ◽  
Blood Cells ◽  
Synthetic Peptide ◽  
Epitope Mapping ◽  
Protein Band ◽  
Band 3 ◽  
Anion Transport ◽  
Transport Protein ◽  
Amino Acid Residues

SUMMARYEpitope mapping of a murine monoclonal antibody (mAb), 5H12, prepared against livePlasmodium falciparum-intected red blood cells indicated that the epitope consisted of amino acid residues 474–487 of the human anion transport protein, band 3. mAb 5H12 enhanced cytoadherence, but inhibited the CD36-like mediated resetting. A synthetic peptide based on the sequence of the epitope (FSFCETNGLE) blocked both resetting and cytoadherence, suggesting that this amino acid sequence may form the CD36-like receptor. The CD36-like region of band 3 was antigenically distinct from platelet or endothelial CD36.

scholarly journals 468 AMINO ACID RESIDUES 93–96 OF THE HUMAN GROWTH HORMONE MOLECULE ARE IMPORTANT FOR POLYCLONAL ANTISERUM RECOGNITION

1985 ◽  
Vol 19 (4) ◽  
pp. 188A-188A
Author(s):  
Wendy J Maury ◽  
Alan P Rogol ◽  
Thaddeus E Kelly ◽  
Christopher Y Thomas
Keyword(s):  
Growth Hormone ◽  
Amino Acid ◽  
Human Growth Hormone ◽  
Human Growth ◽  
Polyclonal Antiserum ◽  
Amino Acid Residues

scholarly journals Mapping antigenic epitopes of potato virus Y with antibodies affinity-purified by using overlapping synthetic peptides

1994 ◽  
Vol 3 (2) ◽  
pp. 207-211
Author(s):  
Maija Vihanen-Rantanen ◽  
Reijo Sironen ◽  
Matti Vuento
Keyword(s):  
Amino Acid ◽  
Coat Protein ◽  
Amino Acid Sequence ◽  
Potato Virus ◽  
Synthetic Peptides ◽  
Potato Virus Y ◽  
Detection Methods ◽  
Polyclonal Antisera ◽  
Antigenic Epitopes

Synthetic, overlapping peptides representing the entire amino acid sequence of potato virus Y (PVY) coat protein were used to affinity-purify antibodies from polyclonal antisera to PVY. In testing the binding of the purified antibodies to PVY particles, antigenic epitopes were identified. The N-terminal and C-terminal regions of the PVY coat protein were found to contain most of the antigenic epitopes. The results will facilitate the development of detection methods for PVY based on synthetic peptides.

scholarly journals Epitope mapping of a monoclonal antibody to human glutathione transferase P1–1 the binding of which is inhibited by glutathione

1997 ◽  
Vol 321 (2) ◽  
pp. 531-536
Author(s):  
Takenori TAKAHATA ◽  
Shigeki TSUCHIDA ◽  
Masashi OOMURA ◽  
Takashi MATSUMOTO ◽  
Junichi AZUMI ◽  
...  
Keyword(s):  
Amino Acid ◽  
Epitope Mapping ◽  
Glutathione Transferase ◽  
Three Dimensional ◽  
Cysteine Residue ◽  
Cross Reactivity ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Steric Configuration ◽  
Terminal Amino

Although the three-dimensional structure of human glutathione transferase (GST) P1Ő1 crystallized with a GSH analogue has been reported, its structure in the non-complexed form has not been determined. Four monoclonal antibodies to GST P1Ő1 were produced to facilitate structural analysis. Of these, one, clone d-1 of IgG2a isotype, dose-dependently inhibited the activity of GST P1Ő1 but did not affect the activities of either GST A1Ő1 or M1Ő1. On immunoblotting, the antibody reacted strongly with GST P1Ő1 and weakly with rat GST-P and mouse GST-II, indicating cross-reactivity with Pi-class forms but preferential reactivity with GST P1Ő1. When GST P1Ő1 and the antibody were incubated in the presence of 60 ƁM GSH, no inhibition of activity was found, whereas 1-chloro-2,4-dinitrobenzene had no effect at concentrations up to 10 ƁM. The binding of GST P1Ő1 to antibody adsorbed to Protein AŐSepharose was also prevented by both 0.1 mM GSH and N-ethylmaleimide treatment. Trypsin digests of GST P1Ő1 were resolved by HPLC and a peptide that reacted with the antibody was detected by absorption experiments. N-Terminal amino acid sequencing revealed the peptide to be in the C-terminal portion of the enzyme, stretching from amino acid residues 198 to 208. A synthetic peptide of this sequence also absorbed the antibody. These results suggest that both GSH bound to the active site and N-ethylmaleimide bound to the cysteine residue repress antibody binding to the C-terminal region. Thus this antibody may be useful for examining the steric configuration of the C-terminal and other regions of GST P1Ő1 in the absence of GSH.

scholarly journals Molecular cloning of a cDNA coding for mouse liver xanthine dehydrogenase. Regulation of its transcript by interferons in vivo

1992 ◽  
Vol 283 (3) ◽  
pp. 863-870 ◽  
Author(s):  
M Terao ◽  
G Cazzaniga ◽  
P Ghezzi ◽  
M Bianchi ◽  
F Falciani ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mouse Liver ◽  
De Novo ◽  
Xanthine Dehydrogenase ◽  
Amino Acid Residues ◽  
Cross Hybridization ◽  
Reading Frame ◽  
Ifn Alpha ◽  
High Conservation

The cDNA coding for xanthine dehydrogenase (XD) is isolated from mouse liver mRNA by cross-hybridization with a DNA fragment of the Drosophila melanogaster homologue. Two lambda bacteriophage overlapping clones represent the copy of a 4538-nucleotide-residue-long transcript with an open reading frame of 4005 nucleotide residues, coding for a putative polypeptide of 1335 amino acid residues. Comparison of the deduced amino acid sequence of the mouse XD with those of the Drosophila and the rat homologues shows a high conservation of this protein (55% identity between mouse and Drosophila, and 94% identity between mouse and rat). RNA blotting analysis demonstrates that interferon-alpha (IFN-alpha) and its inducers, i.e. poly(I).poly(C), bacterial lipopolysaccharide (LPS) and tilorone (2,7-bis-[2-(diethylamino)ethoxy]fluoren-9-one), increase the expression of XD mRNA in liver. Poly(I).poly(C) also induces XD mRNA in several other tissues in vivo. Protein synthesis de novo is not required for the elevation of XD mRNA after IFN-alpha treatment, since cycloheximide does not block the induction. The elevation of XD mRNA concentration is relatively fast and precedes the induction of both XD and xanthine oxidase (XO) enzymic activities.

Amino Acid Regions 572–579 and 657–666 of the Spacer Domain of ADAMTS13 Provide a Common Antigenic Core Required for Binding of Antibodies in Patients with Acquired TTP.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 59-59
Author(s):  
Brenda M. Luken ◽  
Ellen A.M. Turenhout ◽  
Paul H.P. Kaijen ◽  
Rob Fijnheer ◽  
Jan Voorberg
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Antigenic Determinant ◽  
Epitope Mapping ◽  
Core Region ◽  
Homologous Sequence ◽  
Amino Acid Residues ◽  
Thrombocytopenic Purpura ◽  
Recombinant Fragment ◽  
Inhibitory Antibodies

Abstract Inhibitory antibodies against ADAMTS13 have been detected in the majority of patients with acquired thrombotic thrombocytopenic purpura. Epitope-mapping studies revealed that antibodies binding to the cysteine-rich/spacer domains of ADAMTS13 are present in plasma of all patients analyzed so far. We have previously shown that a major antigenic determinant located within the spacer domain. To confirm these results we constructed a recombinant fragment consisting of the disintegrin/TSR1/cysteine-rich domains of ADAMTS13 and the spacer domain of ADAMTS1. The resulting ADAMTS13/ADAMTS1 chimera did not react with IgG present in plasma of a panel of patients with acquired TTP. To identify the amino acid residues that are involved in binding of anti-ADAMTS13 antibodies, we constructed a series of 15 hybrids (designated A–O) in which 5–10 amino acids of the ADAMTS13 spacer were exchanged for the homologous sequence of ADAMTS1. Plasma from 6 patients with antibodies directed against the spacer domain was analyzed for reactivity with the ADAMTS13/ADAMTS1 chimeras. Exchange of residues 572–579 (hybrid C) and 657–666 (hybrid M) of ADAMTS13 for the corresponding sequence of ADAMTS1 completely abolished the binding of antibodies from all 6 patients. Other regions of the ADAMTS13 spacer were also involved in binding of antibodies from patient plasma. Regions 580–587 (D), 602–620 (G,H), 629–638 (J), and 667–767 (N) of the ADAMTS13 spacer domain contributed to binding of antibodies from patients 2, 4, and 5 (epitope present within regions CDGHJMN). IgG derived from patient 1 required region 602–620 (G,H) for binding to the ADAMTS13 spacer (CGHM-epitope). For antibodies of patient 3, residues 564–571 (B), 580–587 (D), and 629–638 (J) were required (BCDJM-epitope), whereas replacement of residues 602–610 (G) and 629–638 (J) greatly diminished binding of antibodies derived from patient 6 (CGJM-epitope). Despite the presumably polyclonal origin of the antibodies present in plasma of the patients, our results suggest that residues 572–579 (C) and 657–666 (M) comprise a common antigenic core region within the spacer domain that is crucial for binding of anti-ADAMTS13 antibodies in all six patients. Amino acid residues derived from other regions that spatially surround this common antigenic core further modulate binding of antibodies to the spacer domain.

scholarly journals Amino Acids in Positions 48, 52, and 73 Differentiate the Substrate Specificities of the Highly Homologous Chlorocatechol 1,2-Dioxygenases CbnA and TcbC

2005 ◽  
Vol 187 (15) ◽  
pp. 5427-5436 ◽  
Author(s):  
Shenghao Liu ◽  
Naoto Ogawa ◽  
Toshiya Senda ◽  
Akira Hasebe ◽  
Kiyotaka Miyashita
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Ralstonia Eutropha ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Sequence Comparisons ◽  
Substrate Specificities ◽  
High Conservation ◽  
The Difference

ABSTRACT Chlorocatechol 1,2-dioxygenase (CCD) is the first-step enzyme of the chlorocatechol ortho-cleavage pathway, which plays a central role in the degradation of various chloroaromatic compounds. Two CCDs, CbnA from the 3-chlorobenzoate-degrader Ralstonia eutropha NH9 and TcbC from the 1,2,4-trichlorobenzene-degrader Pseudomonas sp. strain P51, are highly homologous, having only 12 different amino acid residues out of identical lengths of 251 amino acids. But CbnA and TcbC are different in substrate specificities against dichlorocatechols, favoring 3,5-dichlorocatechol (3,5-DC) and 3,4-dichlorocatechol (3,4-DC), respectively. A study of chimeric mutants constructed from the two CCDs indicated that the N-terminal parts of the enzymes were responsible for the difference in the substrate specificities. Site-directed mutagenesis studies further identified the amino acid in position 48 (Leu in CbnA and Val in TcbC) as critical in differentiating the substrate specificities of the enzymes, which agreed well with molecular modeling of the two enzymes. Mutagenesis studies also demonstrated that Ile-73 of CbnA and Ala-52 of TcbC were important for their high levels of activity towards 3,5-DC and 3,4-DC, respectively. The importance of Ile-73 for 3,5-DC specificity determination was also shown with other CCDs such as TfdC from Burkholderia sp. NK8 and TfdC from Alcaligenes sp. CSV90 (identical to TfdC from R. eutropha JMP134), which convert 3,5-DC preferentially. Together with amino acid sequence comparisons indicating high conservation of Leu-48 and Ile-73 among CCDs, these results suggested that TcbC of strain P51 had diverged from other CCDs to be adapted to conversion of 3,4-DC.

scholarly journals Antigenica and Genetic Characterization of Identified Rotavirus Strains in Qatar in Response to Rotarix Vaccine Usage

2020 ◽  
Author(s):  
Shilu M. Mathew ◽  
Malak Ibrahim ◽  
Asmaa Al Thani ◽  
Khalid Al Ansari ◽  
Hassan Zaraket ◽  
...  
Keyword(s):  
Amino Acid ◽  
Antigenic Variation ◽  
Phylogenetic Analyses ◽  
Amino Acid Residues ◽  
Exact Test ◽  
Number Of Children ◽  
Rotavirus Vaccines ◽  
Antigenic Domains ◽  
Antigenic Epitopes

To identify genetic and antigenic variation in RV in response to vaccine usage. Methods: A total of 231 RV-positive fecal samples were collected from children suffering from AGE during three-year study period between June 2016 and June 2019. The age of the subjects ranged between 2 months and 14 years (median of 16 months). RV genotyping and neutralizing regions, which include both VP4 (Ptype) and VP7 (G type), were amplified and sequenced. We characterized amino acid sequence variability and predicted antigenicity compared to the Rotarix vaccine strain. Phylogenetic analyses were performed using MEGA7.0. Fisher’s exact test was used to run the statistical analysis for the clinical and demographical characteristics of circulating strains. Results: RV infection was most common in children between 3-36 months of age. Among the RV-positive cases, 135 (59.3%) had been vaccinated using either of the RV vaccines available. The number of children vaccinated with one and two-dose was 53 (39.2%) and 82 (60.8%), respectively. The percentage reduction of disease in a vaccinated group of pediatrics compared to an unvaccinated group of pediatrics was 25%. Of these, 108 (78.2%) experienced diarrhea for less than three days, and only eight (6.7%) had diarrhea for more than five days. All vaccinated children showed mild to moderate dehydration except for ten children who were then treated with intravenous fluids. G3 strains were the most strains detected (40%) followed by G2 (17.7%), G4 (16.8%), G9 (15%), G1 (9%), and G8 (0.9%). The dominant RV strains during the study period were G3P [8] (30.8%), G2P [8] (12.3%), G4P [8] (11.7%) and G1P[8] (10.4%). Comparisons of the amino acid residues defining the VP7 and VP4 antigenic domains revealed several mismatches between G1P [8] strains and the G1 and P [8] strains contained in the currently licensed rotavirus vaccines Rotarix. Eighty percent (n=8) of the G1 genotype specimens harbored three amino acid substitutions (N94S, S123N, and M217T) in 7‐ 1a and 7‐ 2b antigenic sites in comparison to the Rotarix vaccine. The P [8] strains with G4 and G9 counterparts showed the highest degree of variation among all specimens with known G genotype. These viruses had 15 and 13 substitutions in their VP4 antigenic epitopes when compared with the P [8] component of the Rotarix vaccines. Conclusion: This study suggests genetic variability in G1 genotype specimens to escape the vaccine-derived immune response. It also identified the wide diversity of circulating RV genotypes in Qatar.

scholarly journals Cloning and characterization of the cDNA coding for a polymyositis-scleroderma overlap syndrome-related nucleolar 100-kD protein.

1992 ◽  
Vol 176 (4) ◽  
pp. 973-980 ◽  
Author(s):  
M Blüthner ◽  
F A Bautz
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Sequence ◽  
Epitope Mapping ◽  
Consensus Sequence ◽  
Overlap Syndrome ◽  
Amino Acid Residues ◽  
Protein Sequence Analysis ◽  
Protein Fragment

About 50% of patients with the polymyositis-scleroderma overlap syndrome are reported to have autoantibodies to a nucleolar particle termed PM/Scl. The particle consists of several polypeptides of which two proteins of 75 and 100 kD have been identified as the major antigenic components. Here we report on the cDNA cloning and partial epitope mapping of the 100-kD autoantigen from human placenta and HeLa lambda gt11 libraries. The deduced amino acid sequence encodes a protein of 885 amino acid residues with a molecular mass of 100.8 kD. Rabbit antibodies raised against a recombinant protein fragment reacted in immunofluorescence and immunoblotting in the same manner as human autoantibodies directed against the nucleolar 100-kD protein. Sequence analysis shows close homology to a consensus sequence of 12 amino acids from serine/threonine kinases, suggesting a possible function for this autoantigen. A major antigenic region is found to be located within the NH2-terminal third of the polypeptide.

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