Purification, characterization and preliminary X-ray diffraction analysis of a cold-active lipase (CpsLip) from the psychrophilic bacterium Colwellia psychrerythraea 34H

被引：11

作者：

Do, Hackwon ^{[1
,2
]}

Lee, Jun Hyuck ^{[1
,2
]}

Kwon, Mi Hyun ^{[1
]}

Song, Hye Eun ^{[1
]}

An, Jun Yop ^{[3
]}

Eom, Soo Hyun ^{[3
]}

Lee, Sung Gu ^{[1
,2
]}

Kim, Hak Jun ^{[1
,2
]}

机构：

[1] Korea Polar Res Inst, Div Polar Life Sci, Inchon 406840, South Korea

[2] Univ Sci & Technol, Dept Polar Sci, Inchon 406840, South Korea

[3] Gwangju Inst Sci & Technol, Sch Life Sci, Kwangju 500712, South Korea

来源：

ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS | 2013年 / 69卷

关键词：

ADAPTATION; SEQUENCE; QUALITY; ENZYMES;

D O I：

10.1107/S1744309113019428

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

The putative lipase CpsLip from the psychrophilic bacterium Colwellia psychrerythraea 34H encodes a 34 538 Da, 308-amino-acid protein. In this study, CpsLip (UniProtKB code Q486T5) was expressed as an N-terminal hexahistidine fusion protein in Escherichia coli and purified by affinity and size-exclusion chromatography. The expression and purification of CpsLip enabled characterization of the lipase enzymatic properties of the protein. The optimal activity temperature and pH of the recombinant protein were 298 K and pH 7, respectively. CpsLip maintained over 80% activity in the low-temperature range (278-288 K), thereby suggesting that CpsLip is a cold-active lipase. Substrate-specificity analysis demonstrated that CpsLip exhibits maximum activity towards the C12 acyl group. In addition, sequence-alignment results revealed that CpsLip has a highly conserved catalytic triad in the active site consisting of residues Ser111, Asp135 and His283. Moreover, purified CpsLip was successfully crystallized using the hanging-drop vapour-diffusion method and a complete diffraction data set was collected to 4.0 angstrom resolution using synchrotron radiation on the BL-5A beamline of the Photon Factory.

引用

页码：920 / 924

页数：5

共 18 条

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