Molecular cloning and characterization of a cDNA for low-temperature inducible cytosolic glucose 6-phosphate dehydrogenase gene from Chlorella vulgaris and expression of the gene in Saccharomyces cerevisiae

A cDNA clone corresponding to the gene for glucose 6-phosphate dehydrogenase (E.C.1.1.1.49) was isolated from a cDNA library constructed from poly(A)+RNA from Chlorella vulgaris C-27, a frost hardy strain. The cDNA clone was designated as Cvcg6pdh. The length of Cvcg6pdh was 1845 bp and the clone coded for 521 amino acids. The deduced amino acid sequence of Cvcg6pdh showed sequence homology to cytosolic G6PDHs from other higher plants rather than chloroplastic ones. Northern blot analysis of a transcript of Cvcg6pdh gene showed that the gene was down-regulated once and then induced after 12-h hardening. Activity staining also showed that the expression pattern of one G6PDH isoform was similar to that of the transcript of the gene. Southern blot analysis of genomic DNA showed that Cvcg6pdh seems to hybridize with, at least, one copy of g6pdh genes. The coding region of the clone was amplified by PCR and the product was introduced into Saccharomyces cerevisiae by using a pTG887 expression vector. The activity of G6PDH in transformed yeast was enhanced up to 8.7 times that of the control strain. Furthermore, after freezing–thawing, the viability of the yeast transformed with pTG887/Cvcg6pdh was significantly higher than that of the control yeast cells carrying pTG887.