Abstract
Several microalgae accumulate high levels of squalene, and as such provide a potentially valuable source of this useful compound. However, the molecular mechanism of squalene biosynthesis in microalgae is still largely unknown. We obtained the sequences of two enzymes involved in squalene synthesis and metabolism, squalene synthase (CrSQS) and squalene epoxidase (CrSQE), from the model green alga Chlamydomonas reinhardtii. CrSQS was functionally characterized by expression in Escherichia coli and CrSQE by complementation of a budding yeast erg1 mutant. Transient expression of CrSQS and CrSQE fused with fluorescent proteins in onion epidermal tissue suggested that both proteins were co-localized in the endoplasmic reticulum. CrSQS-overexpression increased the rate of conversion of 14C-labeled farnesylpyrophosphate into squalene but did not lead to over-accumulation of squalene. Addition of terbinafine caused the accumulation of squalene and suppression of cell survival. On the other hand, in CrSQE-knockdown lines, the expression level of CrSQE was reduced by 59–76% of that in wild-type cells, and significant levels of squalene (0.9–1.1 μg mg–1 cell dry weight) accumulated without any growth inhibition. In co-transformation lines with CrSQS-overexpression and CrSQE-knockdown, the level of squalene was not increased significantly compared with that in solitary CrSQE-knockdown lines. These results indicated that partial knockdown of CrSQE is an effective strategy to increase squalene production in C. reinhardtii cells.
Highlights
Squalene is an important intermediate of sterol biosynthesis in a variety of organisms from bacteria to humans [1]
The above results suggested that the CrSQS and CrSQE genes could encode membrane-bound squalene synthase (SQS) and squalene epoxidase (SQE), respectively, involved in the squalene synthesis and sterol metabolism in C. reinhardtii
We isolated CrSQS and CrSQE, which are responsible for squalene synthesis and metabolism, from the model green alga C. reinhardtii, and demonstrated their enzymatic activities
Summary
Squalene is an important intermediate of sterol biosynthesis in a variety of organisms from bacteria to humans [1]. Squalene has been used as a natural antioxidant, adjuvant for vaccines, dietary supplement and skin moisturizer for therapeutic, pharmacological and cosmetic purposes [1,2]. The main source of squalene is shark liver oil, and alternative sources are desired in order. Characterization of SQS and SQE Genes in Algal Squalene Metabolism
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