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Published on: August 7, 2014
High-molecular-mass isoform of aminopeptidase N/CD13 in serum from cholestatic patients
Makoto Kawai1, Yuya Otake, Yukichi Hara
1Department 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
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.
