Functional Characterization of LpispA, a Key Gene Involved in Carotenoid Biosynthesis in Lactiplantibacillus plantarum
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Abstract:
Carotenoids exhibit various important physiological activities, including anti-oxidation and anti-tumor properties. Compared with plant extracts and chemically synthesized compounds, carotenoids are considered safe for human consumption. Moreover, microbial biosynthesis offers an economically viable approach to the synthesis of carotenoids. As a novel source of the carotenoid C30, the carotenoid biosynthetic pathway in Lactiplantibacillus plantarum has not been fully elucidated. Based on transcriptome analyses performed previously, in this study, the LpispA gene was cloned from a marine L. plantarum strain, and phylogenetic analysis was performed. Subsequently, the cloned open reading frame sequence was expressed in the reconstructed plasmid pAC-BETA∆E-ispA within a functional complementation system using Escherichia coli for carotenoid biosynthesis. In addition, ultraviolet-visible light spectroscopy and reversed-phase high-performance liquid chromatography were employed for component detection of the pigment productions. The results indicated that the L. plantarum LpispA gene exhibited the closest genetic relationship with the corresponding gene from Erwinia persicina. A β-carotene (C40) yield of 0.05 mg/g (dry cell weight) was achieved in the engineered E. coli strain with pAC-BETA∆E-ispA. In conclusion, the bioactivity of geranylgeranyl pyrophosphate synthase expressed by LpispA in the functional complementation E. coli system resulted in the biosynthesis of β-carotene. In other words, LpispA, isolated from L. plantarum, is a multifunctional enzyme in carotenoid biosynthesis, contributing to the biosynthesis of both C30 and C40 carotenoids in bacteria. These results may establish an experimental foundation for investigating carotenoid biosynthesis in L. plantarum and pave the way for potential applications of L. plantarum in the production of carotenoid supplements.
