CLONING AND EXPRESSION OF GENES INVOLVED IN LIPID BIOSYNTHESIS FROM MICROALGAE

Abstract

Production of biofuel directly from CO2 is a challenging approach to solve the near world energy shortage and environmental problems. Accumulating evidence showed that photosynthetic microorganisms would serve as attractive sources because of several unique metabolisms for lipid biosynthesis. Microalgal lipid metabolism is one of the attractive targets to enhance their efficiency in biofuel production by genetic engineering. In the study, caleosin gene encoding lipid droplet protein was successfully isolated from green alga Chlorella vulgaris TISTR 8580 by RACE-method. The deduced polypeptide comprises of 279 amino acid residues with a molecular mass of 31,047 dalton. Chlorella caleosin was expressed and functionally analyzed. It was shown that Chlorella caleosin had no calcium binding property but it had peroxygenase activity. It was found to be a hemoprotein. Semiquantitative RT-PCR revealed that caleosin gene was up-regulated under salt stress and nitrogen deficiency. Furthermore, gene involved in fatty acid biosynthesis was also performed. Acyl Carrier Protein synthetase (ACP) gene from fresh water cyanobacterium Synechococcus elongatus PCC 7942 was cloned and overexpressed under native promoter. Intracellular fatty acid profiles in mock control and overexpressing cells were compared. Palmitic and palmitoleic acids were major fatty acids. Interestingly, total fatty acid of the overexpressing strain was higher than the control approximately 3.7-fold under normal condition but no difference under stress condition was observed. Therefore, metabolic engineering of caleosin or ACP has potential to further enhance biofuel production in microalgae.