Epitope analysis of the major reactive region of the 100-kd protein of PM-Scl autoantigen.
OBJECTIVE
To localize the epitope(s) bound by anti-PM-Scl antibodies in the N-terminal half of the 100-kd protein, the major antigen of the PM-Scl complex.


METHODS
Investigations were performed by immunoblotting 20 anti-PM-Scl positive sera against bacterially expressed, polymerase chain reaction-derived deletion mutants of the S1 fragment (amino acids 11-437), enzyme-linked immunosorbent assay (ELISA) screening against synthesized serial octapeptides, and ELISA screening, with anti-PM-Scl positive sera, against a synthesized 21-amino acid peptide covering the active region.


RESULTS
Anti-PM-Scl positive sera retained full immunoblot activity with fragment 207-436 and most activity with fragment 11-241, but had markedly decreased activity against fragments 236-436 and 11-212, indicating a major epitope in the aa 207-241 region. Fusion proteins with smaller fragments localized this activity between aa 226 and aa 246. Of 42 anti-S1-positive, anti-PM-Scl positive sera tested by ELISA against a synthetic peptide of this region, 36 were definitely positive, 4 borderline, and 2 negative. Similar activity was seen with a peptide from which proline 228 was deleted. Three additional epitope areas were found in S1, but each reacted with only a few sera. Anti-PMScl positive sera did not react with any octapeptide spanning the major epitope area (aa 207-246).


CONCLUSIONS
The main immunoblot epitope of the PM-Scl 100-kd protein is within a central area of 21 aa (aa 226-246), but is longer than the usual linear epitope. This peptide may be useful in patient testing. Three minor epitopes in S1 may also be recognized by some sera.
Ge
Q
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Arthritis/Immunology Section, Oklahoma Medical Research Foundation, Oklahoma City 73104, USA.
Wu
Y
Y
James
J A
JA
Targoff
I N
IN
D000939:Epitopes / Q000032:analysis* / Q000378:metabolism; D004926:Escherichia coli; D005095:Exoribonucleases; D063326:Exosome Multienzyme Ribonuclease Complex; D005786:Gene Expression Regulation; D006801:Humans; D007128:Immunoglobulin Fragments / Q000378:metabolism; D009687:Nuclear Proteins / Q000276:immunology*; D017285:Polymyositis / Q000097:blood / Q000276:immunology*; D012595:Scleroderma, Systemic / Q000097:blood / Q000276:immunology*; D013577:Syndrome
D000939:Epitopes; D007128:Immunoglobulin Fragments; D009687:Nuclear Proteins; D005095:Exoribonucleases; D063326:Exosome Multienzyme Ribonuclease Complex; C077044:EXOSC10 protein, human