Identification of a triacylglycerol lipase in the diatom Phaeodactylum tricornutum

• Published in: Biochimica et biophysica acta 2016-03, Vol.1861 (3), p.239-248
• Main Authors: Barka, Frederik, Angstenberger, Max, Ahrendt, Tilman, Lorenzen, Wolfram, Bode, Helge B., Büchel, Claudia
• Format: Article
• Language: English
• Subjects: Amino Acid Motifs; Amino Acid Sequence; Antisense; Diatoms - enzymology; Diatoms - genetics; Gene Expression Regulation, Enzymologic; Gene Knockdown Techniques; Genotype; Hydrolysis; Kinetics; Lipase; Lipase - chemistry; Lipase - genetics; Lipase - metabolism; Lipids; Molecular Sequence Data; Phenotype; Phylogeny; Polyunsaturated fatty acids; Recombinant Proteins - metabolism; RNA, Antisense - genetics; RNA, Antisense - metabolism; Triglycerides - metabolism
• ISSN: 1388-1981; 0006-3002; 1879-2618
• DOI: 10.1016/j.bbalip.2015.12.023

Diatoms accumulate triacylglycerols (TAGs) as storage lipids, but the knowledge about the molecular mechanisms of lipid metabolism is still sparse. Starting from a partial sequence for a putative TAG-lipase of the diatom Phaeodactylum tricornutum retrieved from the data bases, we have identified the full length coding sequence, tgl1. The gene encodes an 813 amino acid sequence that shows distinct motifs for so called “true” TAG-lipases [EC 3.1.1.3] that have been functionally characterized in model organisms like Arabidopsis thaliana and Saccharomyces cerevisiae. These lipases mediate the first initial step of TAG breakdown from storage lipids. To test whether Tgl1 can act as a TAG-lipase, a His-tagged version was overexpressed in Escherichia coli and the protein indeed showed esterase activity. To identify the TAG degrading function of Tgl1 in P. tricornutum, knock-down mutant strains were created using an antisense RNA approach. In the mutant cell lines the relative tgl1-mRNA-level was reduced up to 20% of that of the wild type, accompanied by a strong increase of TAG in the lipid extracts. In spite of the TAG accumulation, the polar lipid species pattern appeared to be unchanged, confirming the TAG-lipase function of Tgl1. •Tgl1 is a key enzyme of the lipid catabolism of the diatom alga P. tricornutum.•The complete coding sequence of Tgl1 is presented.•Heterologously overexpressed protein shows esterase function.•Knock-down of this enzyme in P. tricornutum leads to TAG accumulation.•Overall fatty acid composition is specifically affected by the mutation.