| Co-Investigator(Kenkyū-buntansha) |
TAJIMA Gen-ichi Tohoku Univ., Grad. Schl. Of Sci., Research Assistant, 大学院・理学研究所, 助手 (00197360)
SHIKAMA Keiji PHP Laboratory, Director, 所長 (40004337)
OCHLAI Takehiko Fukushima Med. Univ., Schl. Of Med., Professor, 医学部, 教授 (40075031)
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| Research Abstract |
A hemoglobin-like protein from yeast, Candida norvegensis, is composed of a single polypeptide chain with 389 amino acid residues, but of two distinct domains carrying different functions: One is a heme-containing oxygen binding domain in the N-terminal region and the other is a FAD containing reductase domain found in the remaining C-terminal region (Iwaasa, H., Takagi, T. and Shikama, K. (1992) J. Mol. Biol. 227, 948-954; Shikama, K., Matsuoka, A. and Iwaasa, H. (1995) Int. J. Biochem. CellBiol. 27, 1107-1115).
To investigate the structure and function of this yeast flavohemoglobin in more detail, we have constructed two expression systems using a vector PMAL-c2 in E. coli cells (TB-1 strain). One is for the whole protein of Candida flavohemoglobin, and the other for the oxygen binding domain (heme domain) alone. The resulting recombinant proteins were then purified using gel filtration on Sephadex G-75 followed by DEAE-cellulose chromatography, and were examined for their spectral and stability properties in veiw of the oxygen-binding function.
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