Proteinase K inactivation of cytosolic aspartate aminotransferase isoenzyme for measurement of human serum mitochondrial aspartate aminotransferase.
We studied a new proteinase K assay method for human serum mitochondrial aspartate aminotransferase. We found that proteinase K showed no inactivation of human mitochondrial aspartate aminotransferase isoenzyme and complete inactivation of cytosolic aspartate aminotransferase. Previous studies have shown that selective proteolytic measurement for mitochondrial aspartate aminotransferase in serum using the protease 401 cleaved peptide bond at Leu 20 from the amino-terminal bond shows complete inactivation of cytosolic aspartate aminotransferase and slight inactivation of mitochondrial aspartate aminotransferase isoenzyme, depending on protease concentration. In this investigation, we found that the proteinase K method does not depend on protease concentration. The proteinase K enzyme inactivation of cytosolic aspartate aminotransferase is caused by the cleavage of the peptide bond at Ileu 21 from the aminoterminal bond. In studies with various animal cytosolic aspartate aminotransferase isoenzymes, proteinase K almost completely inactivated cytosolic aspartate aminotransferase. Precision and correlation using proteinase K for measurement of serum mitochondrial aspartate aminotransferase in human showed a good coefficient of variation (within-run < 4.45%) and a coefficient of correlation of r = 0.985 (N = 125).
Watazu
Y
Y
Department of Laboratory Medicine, Kumamoto University Medical School, Japan.
Uji
Y
Y
Okabe
H
H
Shirahase
Y
Y
Kaneda
N
N
Karmen
A
A
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D001219:Aspartate Aminotransferases / Q000097:blood* / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology; D019286:Endopeptidase K; D006801:Humans; D006868:Hydrolysis; D007527:Isoenzymes / Q000097:blood* / Q000378:metabolism; D008875:Middle Aged; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D012016:Reference Values; D015203:Reproducibility of Results; D012697:Serine Endopeptidases / Q000378:metabolism*
D007527:Isoenzymes; D001219:Aspartate Aminotransferases; D012697:Serine Endopeptidases; D019286:Endopeptidase K