Identification of an aminopeptidase from the skeletal muscle of grass carp (<Emphasis Type="Italic">Ctenopharyngodon idellus</Emphasis>) |
| |
Authors: | Li-Gen Zhou Bing-Xin Liu Le-Chang Sun Kenji Hara Wen-Jin Su Min-Jie Cao |
| |
Institution: | (1) Institute of Food Processing, Zhejiang Academy of Agriculture Sciences, Hangzhou, 310021, China;(2) College of Biological Engineering, The Key Laboratory of Science and Technology for Aquaculture and Food Safety, Jimei University, Jimei, Xiamen, 361021, China;(3) Faculty of Fisheries, Nagasaki University, Nagasaki 852-8521, Japan; |
| |
Abstract: | Aminopeptidases play important roles in turnover of proteins, metabolism of hormones and neurotransmission, cell maturation
and immunological regulations. In the present study, an aminopeptidase was purified to homogeneity from the skeletal muscle
of grass carp by ammonium sulfate fractionation and sequential chromatographic steps, including DEAE-Sephacel, Sephacryl S-200,
hydroxyapatite and Phenyl-Sepharose. The purified enzyme revealed a molecular mass of approximately 105 kDa both on SDS–PAGE
and on gel filtration of Superdex 200. The enzymatic activity toward synthetic substrates was optimal at 40°C and pH 7.0–7.5.
Metal-chelating agents such as EDTA and EGTA effectively inhibited the enzyme activity while inhibitors to serine, asparatic
and cysteine proteinases did not show much effect, suggesting its belonging to metalloproteinase family. A specific aminopeptidase
inhibitor bestatin was most effective in suppressing the enzymatic activity and performed in a competitive fashion. The enzymatic
activity was slightly enhanced by metal ions of Mg2+ and Mn2+ while inhibited to different extents by Co2+, Cu2+, Zn2+ and Ca2+. Sulfhydryl reagent was necessary to maintain its activity. Purified enzyme demonstrated amidolytic activity most effectively
against synthetic aminopeptidase substrate Leu-methylcoumarylamide (MCA) while N-terminal-blocked substrates and myofibrillar
proteins were not hydrolyzed. The enzyme purified in the present study was quite possibly a leucine aminopeptidase (LAP) and
functions during muscular protein metabolism. |
| |
Keywords: | |
本文献已被 SpringerLink 等数据库收录! |
|