High-molecular-mass isoform of aminopeptidase N/CD13 in serum from cholestatic patients.
BACKGROUND
Because non-denaturing electrophoresis and aminopeptidase activity staining often detect noncovalent multi-enzyme complexes, we adopted procedures to specifically detect the aminopeptidase N (APN) molecule itself in liver disease serum.


METHODS
Sera or their immunoprecipitate with anti-APN monoclonal antibody were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or two-dimensional electrophoresis and subsequent Western blotting with rabbit anti-APN serum.


RESULTS
In all the patient sera examined, the 140-kDa APN isoform was predominant. In all the sera from 10 patients with cholestatic diseases (8 with extra-hepatic cholestasis and 2 with primary biliary cirrhosis), we observed the 260-kDa isoform that was immunoprecipitated with monoclonal APN antibodies and had a similar isoelectric point to the 140-kDa isoform. However, the 260-kDa isoform was observed faintly in 2 out of 12 patients with other liver diseases, including chronic hepatitis and cirrhosis.


CONCLUSIONS
We found a novel high-molecular-mass APN isoform (260-kDa) in serum, which is highly likely to be a homodimer of APNs bound covalently and a promising marker of cholestasis. This suggests increased cross-linking reaction between two APN molecules in cholestatic patients.
Kawai
Makoto
M
Department of Biochemistry and Biophysics, Graduate School of Allied Health Sciences, Tokyo Medical and Dental University, 5-45, Yushima 1-chome, Bunkyo-ku, Tokyo 113-8519, Japan. m.kawai.mtec@tmd.ac.jp
Otake
Yuya
Y
Hara
Yukichi
Y
D000626:Aminopeptidases / Q000097:blood* / Q000235:genetics / Q000276:immunology; D015153:Blotting, Western; D002779:Cholestasis / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006801:Humans; D007526:Isoelectric Point; D007527:Isoenzymes / Q000097:blood / Q000235:genetics / Q000276:immunology; D008970:Molecular Weight; D011233:Precipitin Tests; D012680:Sensitivity and Specificity
D007527:Isoenzymes; D000626:Aminopeptidases