Purification and characterization of trypsin from the pyloric ceca of orange-spotted grouper, Epinephelus coioides

in Sciences Citation Index(SCI), 科學引文索引資料庫(SCI)
標題Purification and characterization of trypsin from the pyloric ceca of orange-spotted grouper, Epinephelus coioides
出版類型SCI(Sciences Citation Index)
出版年度2012
AuthorsChun-Hung Liu, 劉俊宏, Ya-Li Shiu 徐雅麗, & Liao Ming-Huei 徐睿良
開始頁837
頁數11
出版日期2012 / 6
其他編號0000
中文摘要

Trypsin from the pyloric ceca of orange-spotted grouper, Epinephelus coioides, was purified by fractionation with ammonium sulfate, ionic exchange, and affinity chromatography. The protein was purified 161.85-fold with a yield of 4%. Purified trypsin had an apparent molecular weight of 24 kDa according to an SDS-PAGE analysis. Optimal profiles of temperature and pH of the enzyme were 50°C and 8-10, respectively, using Nα-benzoyl-L: -arginine ethyl ester as the substrate. The results of thermal and pH stability assays showed that the enzyme was stable at temperatures of up to 50°C and in the pH range of 6-8. Trypsin activity decreased with an increasing NaCl concentration (0-0.6 M). The activity of purified trypsin was effectively inhibited by a soybean trypsin inhibitor and N-p-tosyl-L: -lysine chloromethyl ketone, and was slightly inhibited by iodoacetic acid, ethylenediaminetetraacetic acid, 1-(L: -trans-epoxysuccinyl-leucylamino)-4-guanidinobutane, and pepstatin A. Protein identification of the purified protease showed that the sequences of two peptides, LGEHNI and NLDNDIML, were highly homologous to other fish trypsins. The measurement of trypsin activity in different tissues showed that the highest activity was detected in pyloric ceca, followed by anterior intestine, middle intestine, hind intestine and spleen, but very low activities were found in other tissues. An inverse relationship between the trypsin activity in four tissues of pyloric ceca, anterior intestine, middle intestine and hind intestine and fish body weight as a result of increased pepsin in stomach indicated grouper growth status was increased.

期刊名稱Fish Physiology and Biochemistry
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