Document Detail


Purification and characterization of trypsin from the intestine of hybrid tilapia (Oreochromis niloticusxO.aureus).
MedLine Citation:
PMID:  19899756     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Trypsin from the intestine of hybrid tilapia (Oreochromis niloticus x O.aureus) was purified by the following techniques: acetone precipitation, ammonium sulfate fractionation, Sephacryl S-200 gel filtration, and DEAE-sephacel ion exchange chromatography. The purified enzyme was determined to be homogeneous by polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate (SDS)-PAGE. The molecular weight was estimated as 22,000 Da. The optimum pH and temperature of the enzyme for the hydrolysis of casein were determined to be 9.0 and 60 degrees C, respectively. The enzyme was stable over a broad pH range from 7.0 to 12.0 at 30 degrees C, and the enzyme was inactive at temperatures above 50 degrees C. The behavior of the enzyme for the hydrolysis of casein followed Michaelis-Menten kinetics with Km of 0.46 mg/mL. The purified enzyme was inhibited by the general serine protease inhibitor phenyl methyl sulphonyl fluoride (PMSF) and also by the specific trypsin inhibitor N-p-tosyl-L-lysine chloromethyl ketone (TLCK) using Nalpha-CBZ-L-lysine p-nitrophenyl ester hydrochloride (CBZ-Lys.pNP) as a substrate. The protease was inhibited by the following ions in decreasing order: Zn2+>Fe3+>Cu2+>Al3+>Co2+=Pb2+>Cd2+>Mn2+. The ions Li+, Na+, K+, Mg2+, and Ba2+ had little effect on the enzyme, and Ca2+ can partially promote its activity at low concentration.
Authors:
Qin Wang; Zhi-Xing Gao; Nong Zhang; Yan Shi; Xiao-Lan Xie; Qing-Xi Chen
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of agricultural and food chemistry     Volume:  58     ISSN:  1520-5118     ISO Abbreviation:  J. Agric. Food Chem.     Publication Date:  2010 Jan 
Date Detail:
Created Date:  2010-01-06     Completed Date:  2010-03-22     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0374755     Medline TA:  J Agric Food Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  655-9     Citation Subset:  IM    
Affiliation:
Key Laboratory of Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Science, Xiamen University, Xiamen 361005, China. qwang@xmu.edu.cn
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MeSH Terms
Descriptor/Qualifier:
Animals
Enzyme Stability
Fish Proteins / chemistry*,  isolation & purification*
Intestines / chemistry,  enzymology*
Kinetics
Molecular Weight
Substrate Specificity
Tilapia / metabolism*
Trypsin / chemistry*,  isolation & purification*
Chemical
Reg. No./Substance:
0/Fish Proteins; EC 3.4.21.4/Trypsin

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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