LOCALIZATION OF A PERNICIOUS-ANEMIA AUTOANTIBODY EPITOPE ON THE ALPHA-SUBUNIT OF HUMAN H,K-ADENOSINE TRIPHOSPHATASE

Citation
Yh. Song et al., LOCALIZATION OF A PERNICIOUS-ANEMIA AUTOANTIBODY EPITOPE ON THE ALPHA-SUBUNIT OF HUMAN H,K-ADENOSINE TRIPHOSPHATASE, Scandinavian journal of gastroenterology, 29(2), 1994, pp. 122-127
Citations number
30
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Gastroenterology & Hepatology
ISSN journal
0036-5521
Volume
29
Issue
2
Year of publication
1994
Pages
122 - 127
Database
ISI
SICI code
0036-5521(1994)29:2<122:LOAPAE>2.0.ZU;2-8
Abstract
Four cDNA fragments encoding different portions of the Lu-subunit of h uman H,K-adenosine triphosphatase (ATPase) were amplified by means of the polymerase chain reaction technique, ligated into the plasmid pGEX -2T, and expressed as glutathione S-transferase fusion proteins in Esc herichia coli. The fragments A (residues 163-313), Ba (residues 360-79 7), Bb (residues 526-797), and C (residues 822-1031) together encompas s 77% of the alpha-subunit and cover most of its cytosolic part. The r eactivities of autoantibodies in the sera from patients with perniciou s anaemia with the recombinant fusion proteins were analysed by immuno blotting. One autoantigenic epitope was found in the NH2-terminal part of the Ba fragment-that is, between residues 360 and 525. No epitope was detected in the other fragments. The Ba fragment was cleaved off f rom the glutathione S-transferase fusion protein by the action of thro mbin and was then further purified. By means of enzyme-linked immunoso rbent assay, 28 of 42 sera (67%) from patients with pernicious anaemia were positive against the purified Ba fragment. The present results p rovide a final proof that the human H,K-ATPase cu-subunit is a major a utoantigen in the parietal cell and that the major epitope is located between residues 360 to 525 on the cytosolic side of the secretory mem brane.