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Purification and biochemical characterization of a puromycin-sensitive aminopeptidase from black carp muscle

Paper ID Volume ID Publish Year Pages File Format Full-Text
34352 45019 2015 7 PDF Available
Title
Purification and biochemical characterization of a puromycin-sensitive aminopeptidase from black carp muscle
Abstract

•An aminopeptidase was purified and characterized from black carp muscle.•The 100-kDa aminopeptidase preferentially hydrolyzed the substrate Lys-MCA.•The aminopeptidase(s) activity decreased slowly in salted fish muscle at 4 °C.•It is noteworthy that aminopeptidase(s) contribute to free amino acids formation in salted fish.

Exopeptidases such as aminopeptidases and carboxypeptidases are believed to contribute to the formation of free amino acids during postmortem storage and the processing of meat. In order to understand the role of aminopeptidases in the generation of amino acids and formation of flavor compounds in Chinese traditional salted fish, an aminopeptidase was purified and characterized from black carp muscle. The peptide mass fingerprinting of this enzyme suggested that it was a puromycin-sensitive aminopeptidase purified to homogeneity by ammonium sulfate fractionation and three chromatographies. Puromycin was further confirmed as a competitive inhibitor with Ki value of 0.25 μM. The 100-kDa enzyme preferentially hydrolyzed substrate Lys-MCA with optimum temperature and pH at 40 °C and 7.5, respectively. At the concentration of 3.2% NaCl and 5% ethanol, the enzyme remained 58.5% and 87.5% of its initial activity, respectively. The aminopeptidase(s) activity decreased slowly and remained 41.1% of its initial activity even after 12 days salting of black carp muscle. These results suggest the possible contribution of fish aminopeptidases to free amino acid formation and flavor generation in Chinese traditional salted fish.

Graphical abstract(A) Changes of potential aminopeptidases activities in black carp muscle during salting from 0 to 12 days. (B) SDS-PAGE. Lane M, molecular weight marker; Lanes 1–5, protein samples from the purification stages of crude extract, 30–70% ammonium sulfate precipitation, DEAE-Sephacel, Phenyl-Sepharose, and Hydroxylapatite. SDS-PAGE and MALDI-TOF mass analysis of aminopeptidase from black carp muscle. (C) MALDI-TOF mass analysis of the purified aminopeptidase.Figure optionsDownload full-size imageDownload as PowerPoint slide

Keywords
Black carp; Salted fish; Aminopeptidase; Purification; Biochemical characterization
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Purification and biochemical characterization of a puromycin-sensitive aminopeptidase from black carp muscle
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 50, Issue 7, July 2015, Pages 1061–1067
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us