Expression and characterization of a novel enantioselective lipase from Aspergillus fumigatus.
A 1,080-bp cDNA (CGMCC 2873) encoding of a cold-active lipase of Aspergillus fumigatus (AFL67) was cloned and expressed in Escherichia coli for the first time. The new lipase, AFL67, was one-step purified by 8.30 folds through Ni-NTA affinity chromatography with a recovery of 86.8 %. The specific activity of purified AFL67 was 449 U mg(-1) on p-NP hexanoate. AFL67 preferentially hydrolyzed p-nitrophenyl esters of short- and medium-chain fatty acids, with p-nitrophenyl hexanoate the maximum. The optimum temperature and pH was 15 °C and 7.5, respectively. The purified AFL67 was stable at 10-25 °C for 30 min, and in the pH range of 6.0-9.0 for 16 h (at 4 °C). Its activity was increased by 47 and 50 %, in the presence of 10 % (v/v) ethanol and isopropanol, respectively. The new lipase AFL67 highly enantioselectively deacylated (S)-α-acetoxyphenylacetic acid (APA) and o-Cl-APA, m-Cl-APA, and p-Cl-APA to (S)-mandelic acid and its derivates. These features render this cold-active novel lipase AFL67 attractive for biotechnological applications in the field of enantioselective synthesis of chiral mandelic acids, o-acylated mandelic acids, and their derivates and detergent additives.
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, People's Republic of China.
Jiao-Jiao
JJ
Shangguan
Li-Qiang
LQ
Fan
Xin
X
Ju
Qing-Qing
QQ
Zhu
Fu-Jun
FJ
Wang
Jian
J
Zhao
Jian-He
JH
Xu
D000085:Acetates / Q000737:chemistry / Q000378:metabolism; D001232:Aspergillus fumigatus / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004795:Enzyme Stability / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D008333:Mandelic Acids / Q000737:chemistry / Q000378:metabolism; D008670:Metals / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D011480:Protease Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012997:Solvents / Q000494:pharmacology; D013237:Stereoisomerism; D013379:Substrate Specificity; D013696:Temperature
D000085:Acetates; D008333:Mandelic Acids; D008670:Metals; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D012997:Solvents; D008049:Lipase; C037938:mandelic acid