Identification of a malonyl CoA-acyl carrier protein transacylase and its regulatory role in fatty acid biosynthesis in oleaginous microalga Nannochloropsis oceanica.

Oleaginous microalgae hold great promises for biofuel production. However, commercialization of microalgal biofuels remains impracticable due to the lack of suitable industrial strains with high growth rate and lipid productivity. Engineering of metabolic pathways is a potential strategy for the improvement of microalgal strains for the production of lipids and also value-added products in microalgae. Malonyl CoA-acyl carrier protein transacylase (MCAT) has been reported to be involved in fatty acid biosynthesis. Here, we identified a putative MCAT in the oleaginous marine microalga Nannochloropsis oceanica. NoMCAT overexpressing N. oceanica showed a higher growth rate and photosynthetic efficiency. The neutral lipid content of engineered lines showed a significant increase by up to 31% compared to wild type. Gas chromatography-mass spectrometry analysis revealed that NoMCAT overexpression significantly altered the fatty acid composition. The composition of eicosapentaenoic acid (C20:5), which is a polyunsaturated fatty acid necessary for animal nutrition, increased by 8%. These results demonstrate the role of MCAT in enhancing fatty acid biosynthesis and growth in microalgae, and also provide an insight into metabolic engineering of microalgae with high industrial potential.

Purification and characterization of the Mycobacterium tuberculosis FabD2, a novel malonyl-CoA:AcpM transacylase of fatty acid synthase.

Malonyl coenzyme A (CoA)-acyl carrier protein (ACP) transacylase (MCAT) is an essential enzyme in fatty acid and mycolic acid biosynthesis of Mycobacterium tuberculosis. fabd2 is a novel gene coding MCAT in M. tuberculosis besides another known fabd. In our study, fabd2 was inserted into a bacterial expression vector pET28a resulting in a 6x Histidine-tag fabd2 fusion gene construction. The protein was purified by nickel affinity chromatography and the characterizations of FabD2 have been investigated. The molecular weight of FabD2 was estimated to be 26 kDa by MALDI-TOF. Consistent with the biosynthesis specialty of reported MCATs, FabD2 resulted in a typical activity of bacterial MCATs, which catalyzes the transacylation of malonate from malonyl-CoA to activated holo-ACP. Some physical and chemical differences between FabD2 and FabD also have been found. FabD2 shows dissimilarity with FabD in secondary structure in different pH buffer and MCAT genes RT-PCR results reveal different transcript condition with each other. Furthermore, FabD2 shows low similarity in protein sequence when alignment with other MCATs.