Linked Life Data

19940341

Source:http://linkedlifedata.com/resource/pubmed/id/19940341

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2009-11-26
pubmed:abstractText
The protein termed RA33 was determined to be one major autoantigen in rheumatoid arthritis (RA) patients and antiRA33 auto-antibodies were found to appear shortly after onset of RA. They are often detectable before a final diagnosis can be made in the clinic. The aim of our study is to characterise the epitope of a monoclonal antiRA33 antibody on recombinant RA33 using mass spectrometric epitope mapping. Recombinant RA33 has been subjected to BrCN cleavage and fragments were separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Subsequent in-gel proteolytic digestion and mass spectrometric analysis determined the partial sequences in the protein bands. Western blotting of SDS-PAGE-separated protein fragments revealed immuno-positive, i.e. epitope-containing bands. BrCN-derived RA33 fragments were also separated by high- performance liquid chromatography (HPLC) and immuno-reactivity of peptides was measured by dot-blot analysis with the individual HPLC fractions after partial amino acid sequences were determined. The epitope region identified herewith was compared to data from peptide chip analysis with 15-meric synthetic peptides attached to a glass surface. Results from all three analyses consistently showed that the epitope of the monoclonal antiRA33 antibody is located in the aa79-84 region on recombinant RA33; the epitope sequence is MAARPHSIDGRVVEP. Sequence comparisons of the 15 best scoring peptides from the peptide chip analysis revealed that the epitope can be separated into two adjacent binding parts. The N-terminal binding parts comprise the amino acid residues "DGR", resembling the general physico-chemical properties "acidic/polar-small-basic". The C-terminal binding parts contain the amino acid residues "VVE", with the motif "hydrophobic-gap-acidic". The matching epitope region that emerged from our analysis on both the full-length protein and the 15-meric surface bound peptides suggests that peptide chips are indeed suitable tools for screening patterns of autoantibodies in patients suffering from autoimmune diseases.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
1469-0667
pubmed:author
pubmed:issnType
Print
pubmed:volume
15
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
747-59
pubmed:meshHeading
pubmed-meshheading:19940341-Amino Acid Sequence, pubmed-meshheading:19940341-Antibodies, Monoclonal, pubmed-meshheading:19940341-Arthritis, Rheumatoid, pubmed-meshheading:19940341-Autoantigens, pubmed-meshheading:19940341-Blotting, Western, pubmed-meshheading:19940341-Chromatography, High Pressure Liquid, pubmed-meshheading:19940341-Cyanogen Bromide, pubmed-meshheading:19940341-Electrophoresis, Polyacrylamide Gel, pubmed-meshheading:19940341-Epitope Mapping, pubmed-meshheading:19940341-Heterogeneous-Nuclear Ribonucleoprotein Group A-B, pubmed-meshheading:19940341-Humans, pubmed-meshheading:19940341-Molecular Sequence Data, pubmed-meshheading:19940341-Protein Array Analysis, pubmed-meshheading:19940341-Protein Structure, Quaternary, pubmed-meshheading:19940341-Protein Structure, Tertiary, pubmed-meshheading:19940341-Spectrometry, Mass, Matrix-Assisted Laser...
pubmed:year
2009
pubmed:articleTitle
Mass spectrometric and peptide chip epitope mapping of rheumatoid arthritis autoantigen RA33.
pubmed:affiliation
Proteome Center Rostock, University of Rostock, Schillingallee 69, 18057 Rostock, Germany.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't