Research Article

Molecular cloning and characterization of a tocopherol cyclase gene from Lactuca sativa (Asteraceae)

Published: April 19, 2011
Genet. Mol. Res. 10 (2) : 693-702 DOI: 10.4238/vol10-2gmr1061

Abstract

Tocopherol cyclase is a rate-limiting enzyme involved in tocopherol biosynthesis. The full-length cDNA encoding tocopherol cyclase (designated as LsTC) was cloned from lettuce (Lactuca sativa) for the first time by rapid amplification of cDNA ends (RACE) and characterized by means of quantitative RT-PCR. The full-length cDNA of LsTC was 1675 bp, with an open reading frame of 1521 bp, encoding a tocopherol cyclase protein of 506 amino acids, with a calculated molecular mass of 56.76 kD and an isoelectric point of 6.49. Comparative analysis revealed that LsTC has a close similarity with tocopherol cyclases from other plant species. Bioinformatic analysis indicated that LsTC shares a common evolutionary origin based on sequence and has the closest relationship to tocopherol cyclase from Helianthus annuus. Quantitative RT-PCR analysis suggested that expression of LsTC is induced and strengthened by oxidative stresses, such as strong light and drought. This cloning and characterization of LsTC will be helpful for further understanding of its role in the tocopherol biosynthesis pathway and provide a candidate gene for metabolic engineering of vitamin E.

Tocopherol cyclase is a rate-limiting enzyme involved in tocopherol biosynthesis. The full-length cDNA encoding tocopherol cyclase (designated as LsTC) was cloned from lettuce (Lactuca sativa) for the first time by rapid amplification of cDNA ends (RACE) and characterized by means of quantitative RT-PCR. The full-length cDNA of LsTC was 1675 bp, with an open reading frame of 1521 bp, encoding a tocopherol cyclase protein of 506 amino acids, with a calculated molecular mass of 56.76 kD and an isoelectric point of 6.49. Comparative analysis revealed that LsTC has a close similarity with tocopherol cyclases from other plant species. Bioinformatic analysis indicated that LsTC shares a common evolutionary origin based on sequence and has the closest relationship to tocopherol cyclase from Helianthus annuus. Quantitative RT-PCR analysis suggested that expression of LsTC is induced and strengthened by oxidative stresses, such as strong light and drought. This cloning and characterization of LsTC will be helpful for further understanding of its role in the tocopherol biosynthesis pathway and provide a candidate gene for metabolic engineering of vitamin E.

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