Identification and functional expression of a Delta9-fatty acid desaturase from Psychrobacter urativorans in Escherichia coli.
The Delta9-fatty acid desaturase is a key enzyme in the synthesis of unsaturated fatty acids. The fatty acid composition of membrane phospholipids in Psychrobacter urativorans is characterized by a high degree of desaturation at Delta9 position. Based on CODEHOP-mediated PCR strategy, a novel gene designated as PuFAD9, putatively encoding a Delta9-fatty acid desaturase (PuFAD9), was isolated from P. urativorans. The gene consists of 1,455 bp and codes for 484 amino acids. Analysis of the amino acid sequence reveals three histidine clusters and a hydropathy profile, typical for membrane-bound desaturases. Activity of the PuFAD9 protein, recombinantly expressed in Escherichia coli was confirmed by GC-MS analysis of the cellular fatty acid composition. It was found that the ratio between palmitoleic and palmitic acid in E. coli cells heterologously expressing the PuFAD9 gene was significantly affected by IPTG induction and the growth temperature.
Li
Yan
Y
Institute of Technical Biochemistry, University of Stuttgart, Stuttgart 70569, Germany.
Dietrich
Matthias
M
Schmid
Rolf D
RD
He
Bingfang
B
Ouyang
Pingkai
P
Urlacher
Vlada B
VB
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005229:Fatty Acids, Monounsaturated / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D007544:Isopropyl Thiogalactoside / Q000378:metabolism; D008969:Molecular Sequence Data; D019308:Palmitic Acid / Q000378:metabolism; D044096:Psychrobacter / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D013230:Stearoyl-CoA Desaturase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D005229:Fatty Acids, Monounsaturated; D011994:Recombinant Proteins; C008757:palmitoleic acid; D019308:Palmitic Acid; D007544:Isopropyl Thiogalactoside; D013230:Stearoyl-CoA Desaturase