Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/60781
標題: Functional Proteomic Analysis of Rice Bran Esterases/Lipases and Characterization of a Novel Recombinant Esterase
作者: Chuang, Hsu-Han
Chen, Po-Ting
Wang, Wun-Nai
Chen, Yu-Ting
Shaw, Jei-Fu
關鍵字: Rice bran
lipolytic enzyme
esterase/lipase
two-dimensional gel
electrophoresis
alpha/beta-hydrolase fold
cold-adapted lipase
metagenomic library
bacillus-licheniformis
carboxyl esterases
molecular-cloning
gene
cloning
purification
enzymes
family
摘要: An esterase from rice (Oryza sativa) bran was identified on two-dimensional gel using 4-methylumbelliferyl butyrate as a substrate. The esterase cDNA (870 bp) encoded a 289 amino acid protein (designated OsEST-b) and was expressed in Escherichia coli. The molecular weight of recombinant OsEST-b (rOsEST-b) was 27 kDa, as measured by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Biochemical characterization demonstrated that rOsEST-b was active over a broad temperature range (optimum at 60 degrees C) and preferred alkaline conditions (optimum at pH 9.0). The rOsEST-b showed maximum activity toward p-nitayphenyl butyrate (C(4)) among various p-nitrophenyl esters (C(4)-C(18)), indicating that rOsEST-b is an esterase for short-chain fatty acids. The kinetic parameters under optimal conditions were K(m) = 27.03 mu M, k(cat) = 49 s(-1), and k(cat)/K(m) = 1.81 s(-1) mu M(-1). The activity of rOsEST-b was not influenced by ethylenediaminetetraacetic acid, suggesting that it is not a metalloenzyme. The amino acid sequence analysis revealed that OsEST-b had a conserved pentapeptide esterase/lipase motif but that the essential active site serine (GXSXG) was replaced by cysteine (C). These results suggest that OsEST-b is distinct from traditional esterases/lipases and is a novel lipolytic enzyme in rice bran.
URI: http://hdl.handle.net/11455/60781
ISSN: 0021-8561
Appears in Collections:基因體暨生物資訊學研究所

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