Cloning and stress-respondent transcription of 3-ketoacyl-CoA thiolase gene of Isochrysis galbana (Haptophyta)

摘要

Thiolases are crucial for both synthesis and catabolism of fatty acids, which catalyze the reversible thiolytic cleavage of 3-ketoacyl-CoA into acyl-CoA and acetyl-CoA. The thiolase of a unicellular alga Isochrysis galbana (IgKAT) has not been identified yet. In this study, a full-length cDNA of 3-ketoacyl-CoA thiolase gene (IgKAT) was isolated with real-time PCR and rapid amplification of cDNA ends methods. IgKAT was 1,910 bp in length, containing an open reading frame of 1,188 bp, a 5'-terminal untranslated region (UTR) of 160 bp, and a 3'-UTR of 562 bp. The deduced amino acid sequence of IgKAT was 395 amino acids residues in length with a predicted molecular weight of 40.47 kDa and an isoelectric point of 6.53. IgKAT contained highly conserved domains and activity-center forming amino acids residues. In comparison with the normal temperature (25 A degrees C) and original content of sodium nitrate in f/2 medium (75 mg L-1), high temperature (35 A degrees C) and sodium nitrate limitation (0 mg L-1) upregulated the transcription of IgKAT gene, while low temperature (15 A degrees C) and sodium nitrate excess (150 mg L-1) downregulated the transcription of IgKAT gene in general. Our findings provided an understanding about the response of IgKAT to temperature and nutrition nitrate stresses (those deviating from the normal) at transcription level. These findings will aid to our understanding of the gene regulation of beta-oxidation pathway in response to culture conditions of I. galbana.