Enhancement of menadione stress tolerance in yeast by accumulation of hypotaurine and taurine: co-expression of cDNA clones, from Cyprinus carpio, for cysteine dioxygenase and cysteine sulfinate decarboxylase in Saccharomyces cerevisiae.
Taurine is known to function as a protectant against various stresses in animal cells. In order to utilize taurine as a compatible solute for stress tolerance of yeast, isolation of cDNA clones for genes encoding enzymes involved in biosynthesis of taurine was attempted. Two types of cDNA clones corresponding to genes encoding cysteine dioxygenase (CDO1 and CDO2) and a cDNA clone for cysteine sulfinate decarboxylase (CSD) were isolated from Cyprinus carpio. Deduced amino acid sequences of the two CDOs and that of CSD showed high similarity to those of CDOs and those of CSDs from other organisms, respectively. The coding regions of CDO1, CDO2, and CSD were subcloned into an expression vector, pESC-TRP, for Saccharomyces cerevisiae. Furthermore, to enhance the efficiency of synthesis of taurine in S. cerevisiae, a CDO-CSD fusion was designed and expressed. Expression of CDO and CSD proteins, or the CDO-CSD fusion protein was confirmed by Western blot analysis. HPLC analysis showed that the expression of the proteins led to enhancement of the accumulation level of hypotaurine, a precursor of taurine, rather than taurine. The yeast cells expressing corresponding genes showed tolerance to oxidative stress induced by menadione, but not to freezing-thawing stress.
Honjoh
Ken-ichi
K
Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School, Kyushu University, Fukuoka, 812-8581, Japan. honjoh@agr.kyushu-u.ac.jp
Matsuura
Kanae
K
Machida
Takeshi
T
Nishi
Koutarou
K
Nakao
Miki
M
Yano
Tomoki
T
Miyamoto
Takahisa
T
Iio
Masayoshi
M
D000595:Amino Acid Sequence; D000818:Animals; D002262:Carboxy-Lyases / Q000096:biosynthesis* / Q000235:genetics; D002347:Carps / Q000235:genetics*; D002999:Clone Cells; D050558:Cysteine Dioxygenase / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D030541:Databases, Genetic; D005615:Freezing / Q000009:adverse effects; D015723:Gene Library; D007527:Isoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D018384:Oxidative Stress / Q000187:drug effects*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013312:Stress, Physiological; D013481:Superoxides / Q000378:metabolism; D013654:Taurine / Q000031:analogs & derivatives / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D024483:Vitamin K 3 / Q000633:toxicity*
D007527:Isoenzymes; D013481:Superoxides; D013654:Taurine; C003949:hypotaurine; D024483:Vitamin K 3; D050558:Cysteine Dioxygenase; D002262:Carboxy-Lyases; C019972:sulfoalanine decarboxylase