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Identification and Functional Characterization of the Delta 6‐Fatty Acid Desaturase Gene from Thamnidium elegans
Author(s) -
WANG DEPEI,
LI MINGCHUN,
WEI DONGSHENG,
CAI YI,
ZHANG YINGHUI,
XING LAIJUN
Publication year - 2007
Publication title -
journal of eukaryotic microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.067
H-Index - 77
eISSN - 1550-7408
pISSN - 1066-5234
DOI - 10.1111/j.1550-7408.2006.00136.x
Subject(s) - biology , fatty acid desaturase , open reading frame , complementary dna , peptide sequence , rapid amplification of cdna ends , pichia pastoris , biochemistry , microbiology and biotechnology , gene , amino acid , histidine , fatty acid , recombinant dna , polyunsaturated fatty acid
. A cDNA sequence was cloned from the filamentous fungus Thamnidium elegans As3.2806 using reverse transcription polymerase chain reaction (RT‐PCR) and rapid amplification of cDNA ends method (RACE). Sequence analysis indicated that this cDNA sequence has an open reading frame of 1,380 bp, which encodes a 52.4 kDa peptide of 459 amino acids. The designated amino acid sequence has high similarity with that found in fungal delta 6‐fatty acid desaturases: it shows three conserved histidine‐rich motifs and two hydrophobic domains. A cytochrome b 5‐like domain was observed at the N‐terminus. To elucidate the function of this novel putative desaturase, the open reading frame was cloned into the intracellular expression vector pPIC3.5K and the gene was expressed heterologously in Pichia pastoris . Accumulation of γ‐linolenic acid to the level of 6.83% in total fatty acid demonstrated that the deduced amino acid sequence possesses of delta 6‐fatty acid desaturase activity.