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Improved serological detection of rheumatoid arthritis: a highly antigenic mimotope of carbonic anhydrase III selected in a murine model by phage display.

Araujo GR, Vaz ER, Fujimura PT, Fonseca JE, de Lima LM, Canhão H, Venturini G, Cardozo KH, Carvalho VM, Napimoga MH, Goulart LR, Gonçalves J, Ueira-Vieira C - Arthritis Res. Ther. (2015)

Bottom Line: The specificity and sensitivity of anti-M12 antibodies for RA diagnosis were 91 % and 84.3 %, respectively.The M12 peptide was identified as one that mimics a predicted antigenic site of the carbonic anhydrase III (CAIII) protein, a ubiquitous biomarker that has been identified in patients with other diseases.M12 is the first peptide associated with the CAIII protein that may be used as an antigen for antibody detection to aid in RA diagnosis with high sensitivity and specificity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Nanobiotechnology, Institute of Genetics and Biochemistry, Federal University of Uberlândia, Uberlândia, MG, Brazil. galber.araujo@gmail.com.

ABSTRACT

Introduction: Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that affects around 1% of the human population worldwide. RA diagnosis can be difficult as there is no definitive test for its detection. Therefore, the aim of this study was to identify biomarkers that could be used for RA diagnosis.

Methods: Sera from a collagen-induced arthritis mouse model were used to select potential biomarkers for RA diagnosis by phage display technology. In silico and in vitro analyses were performed to characterize and validate the selected peptides. Samples were classified into three groups: RA; two other immune-mediated rheumatic diseases (systemic lupus erythematosus (SLE) and ankylosing spondylitis (AS)); and healthy controls (HC). Enzyme-linked immunosorbent assay (ELISA) was carried out to determine antibody levels, and diagnostic parameters were determined by constructing receiver operating characteristic curves. Mass spectrometry and Western blot were performed to identify the putative autoantigen that was mimicked by a highly reactive mimotope.

Results: After three rounds of selection, 14 clones were obtained and tested for immunoreactivity analysis against sera from RA and HC groups. The phage-fused peptide with the highest immunoreactivity (M12) was synthesized, and was able to efficiently discriminate RA patients from SLE, AS and HCs (p < 0.0001) by ELISA. The specificity and sensitivity of anti-M12 antibodies for RA diagnosis were 91 % and 84.3 %, respectively. The M12 peptide was identified as one that mimics a predicted antigenic site of the carbonic anhydrase III (CAIII) protein, a ubiquitous biomarker that has been identified in patients with other diseases.

Conclusion: M12 is the first peptide associated with the CAIII protein that may be used as an antigen for antibody detection to aid in RA diagnosis with high sensitivity and specificity.

No MeSH data available.


Related in: MedlinePlus

Peptide localization. a Multiple alignments of M12 peptide and deduced amino acid sequences of carbonic anhydrase III from Homo sapiens and Mus musculus. Conserved (stars) and semi-conserved residues (gray) are highlighted. Valine (V) amino acid is star marked because it matches a position between M12 peptide and human CAIII protein. b Model of the three-dimensional structure predicted in the PyMol server software for the human carbonic anhydrase III with M12 peptide localization
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Fig4: Peptide localization. a Multiple alignments of M12 peptide and deduced amino acid sequences of carbonic anhydrase III from Homo sapiens and Mus musculus. Conserved (stars) and semi-conserved residues (gray) are highlighted. Valine (V) amino acid is star marked because it matches a position between M12 peptide and human CAIII protein. b Model of the three-dimensional structure predicted in the PyMol server software for the human carbonic anhydrase III with M12 peptide localization

Mentions: The multiple sequence alignment revealed several homologous sequences between M12 peptide and the CAIII protein sequences from mouse and human. Nine (75 %) conserved or semi-conserved aa residues of the M12 peptide sequence matched a domain of the CAIII protein (Fig. 4a). A three-dimensional structural alignment was performed to predict the putative epitope site of the M12 peptide in the CAIII protein structure [PDB:3UYN], which confirmed its surface exposure and corroborated to the possible antibody-binding region in the external sequences of the predicted protein (Fig. 4b). Five (41.6 %) aa residues (SPVET) of the M12 peptide was matched to an antigenic region of the human CAIII protein (Fig. 5).Fig. 4


Improved serological detection of rheumatoid arthritis: a highly antigenic mimotope of carbonic anhydrase III selected in a murine model by phage display.

Araujo GR, Vaz ER, Fujimura PT, Fonseca JE, de Lima LM, Canhão H, Venturini G, Cardozo KH, Carvalho VM, Napimoga MH, Goulart LR, Gonçalves J, Ueira-Vieira C - Arthritis Res. Ther. (2015)

Peptide localization. a Multiple alignments of M12 peptide and deduced amino acid sequences of carbonic anhydrase III from Homo sapiens and Mus musculus. Conserved (stars) and semi-conserved residues (gray) are highlighted. Valine (V) amino acid is star marked because it matches a position between M12 peptide and human CAIII protein. b Model of the three-dimensional structure predicted in the PyMol server software for the human carbonic anhydrase III with M12 peptide localization
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4493817&req=5

Fig4: Peptide localization. a Multiple alignments of M12 peptide and deduced amino acid sequences of carbonic anhydrase III from Homo sapiens and Mus musculus. Conserved (stars) and semi-conserved residues (gray) are highlighted. Valine (V) amino acid is star marked because it matches a position between M12 peptide and human CAIII protein. b Model of the three-dimensional structure predicted in the PyMol server software for the human carbonic anhydrase III with M12 peptide localization
Mentions: The multiple sequence alignment revealed several homologous sequences between M12 peptide and the CAIII protein sequences from mouse and human. Nine (75 %) conserved or semi-conserved aa residues of the M12 peptide sequence matched a domain of the CAIII protein (Fig. 4a). A three-dimensional structural alignment was performed to predict the putative epitope site of the M12 peptide in the CAIII protein structure [PDB:3UYN], which confirmed its surface exposure and corroborated to the possible antibody-binding region in the external sequences of the predicted protein (Fig. 4b). Five (41.6 %) aa residues (SPVET) of the M12 peptide was matched to an antigenic region of the human CAIII protein (Fig. 5).Fig. 4

Bottom Line: The specificity and sensitivity of anti-M12 antibodies for RA diagnosis were 91 % and 84.3 %, respectively.The M12 peptide was identified as one that mimics a predicted antigenic site of the carbonic anhydrase III (CAIII) protein, a ubiquitous biomarker that has been identified in patients with other diseases.M12 is the first peptide associated with the CAIII protein that may be used as an antigen for antibody detection to aid in RA diagnosis with high sensitivity and specificity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Nanobiotechnology, Institute of Genetics and Biochemistry, Federal University of Uberlândia, Uberlândia, MG, Brazil. galber.araujo@gmail.com.

ABSTRACT

Introduction: Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that affects around 1% of the human population worldwide. RA diagnosis can be difficult as there is no definitive test for its detection. Therefore, the aim of this study was to identify biomarkers that could be used for RA diagnosis.

Methods: Sera from a collagen-induced arthritis mouse model were used to select potential biomarkers for RA diagnosis by phage display technology. In silico and in vitro analyses were performed to characterize and validate the selected peptides. Samples were classified into three groups: RA; two other immune-mediated rheumatic diseases (systemic lupus erythematosus (SLE) and ankylosing spondylitis (AS)); and healthy controls (HC). Enzyme-linked immunosorbent assay (ELISA) was carried out to determine antibody levels, and diagnostic parameters were determined by constructing receiver operating characteristic curves. Mass spectrometry and Western blot were performed to identify the putative autoantigen that was mimicked by a highly reactive mimotope.

Results: After three rounds of selection, 14 clones were obtained and tested for immunoreactivity analysis against sera from RA and HC groups. The phage-fused peptide with the highest immunoreactivity (M12) was synthesized, and was able to efficiently discriminate RA patients from SLE, AS and HCs (p < 0.0001) by ELISA. The specificity and sensitivity of anti-M12 antibodies for RA diagnosis were 91 % and 84.3 %, respectively. The M12 peptide was identified as one that mimics a predicted antigenic site of the carbonic anhydrase III (CAIII) protein, a ubiquitous biomarker that has been identified in patients with other diseases.

Conclusion: M12 is the first peptide associated with the CAIII protein that may be used as an antigen for antibody detection to aid in RA diagnosis with high sensitivity and specificity.

No MeSH data available.


Related in: MedlinePlus