Purification and properties of an aminopeptidase from the mid-gut gland of scallop ( Patinopecten yessoensis)
An aminopeptidase was isolated from the mid-gut gland of Patinopecten yessoensis. The enzyme was purified from an acetone-dried preparation by extracting, ammonium sulfate precipitation, Hi-Load Q column chromatography, isoelectric focusing, and POROS HP2 and HQ column chromatography. The molecular...
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Published in | Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology Vol. 136; no. 4; pp. 935 - 942 |
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Main Authors | , , , , , , |
Format | Journal Article |
Language | English |
Published |
England
Elsevier Inc
01.12.2003
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Subjects | |
Online Access | Get full text |
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Summary: | An aminopeptidase was isolated from the mid-gut gland of
Patinopecten yessoensis. The enzyme was purified from an acetone-dried preparation by extracting, ammonium sulfate precipitation, Hi-Load Q column chromatography, isoelectric focusing, and POROS HP2 and HQ column chromatography. The molecular weight of the enzyme was estimated to be 61 kDa by SDS-polyacrylamide gel electrophoresis and 59 kDa by gel permeation chromatography. The isoelectric point of the enzyme was 5.2 and the optimum pH was 7.0 toward leucine
p-nitroanilide (Leu-
pNA). The enzyme was inhibited by
o-phenanthroline. The activity of the enzyme treated with
o-phenanthroline was completely recovered by adding excess Zn
2+. Relative hydrolysis rates of amino acid-
pNAs and amino acid-4-methylcoumaryl-7-amides (amino acid-MCAs) indicated that the enzyme preferred substrates having Ala or Met as an amino acid residue. The enzyme had a
K
m of 32.2 μM and
k
cat of 29.5 s
−1 with Ala-
pNA and a
K
m of 11.1 μM and
k
cat of 9.49 s
−1 with Ala-MCA. The enzyme sequentially liberated amino acids from the amino-termini of Ala–Phe–Tyr–Glu. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 1096-4959 1879-1107 |
DOI: | 10.1016/j.cbpc.2003.09.008 |