Yeast flavohemoglobin from Candida norvegensis. Its structural, spectral, and stability properties.
Flavohemoglobin was isolated directly from the yeast Candida norvegensis and studied on its structural, spectral, and stability properties. In Candida flavohemoglobin, the 155 N-terminal residues make a heme-containing domain, while the remaining 234 C-terminal residues serve as a FAD-containing reductase domain. A pair of His-95 and Gln-63 was assigned to the proximal and distal residues, respectively. In purification procedure FAD was partially dissociated on a Butyl-Toyopearl column, so that FAD-lacking flavohemoglobin was also obtainable. In this ferric species, the Soret and charge-transfer bands were all characteristic of a penta-coordinate form. Compared with the recombinant heme domain expressed in Escherichia coli, we have measured the autoxidation rate over a wide pH range. The resulting pH dependence curves were then analyzed in terms of a nucleophilic displacement mechanism. As a result, the heme domain was found to be extremely susceptible to autoxidation, its rate being more than 100 times higher than that of sperm whale MbO2. However, this inherently high oxidation rate was dramatically suppressed in Candida flavohemoglobin to an extent almost comparable to the stability of mammalian myoglobins. These new findings lead us to conclude that Candida flavohemoglobin, differently from bacterial flavohemoglobins, can serve as an oxygen storage protein in aerobic conditions.
Kobayashi
Gen
G
Biological Institute, Graduate School of Life Sciences, Tohoku University, Sendai 980-8578, Japan.
Nakamura
Takashi
T
Ohmachi
Hitoshi
H
Matsuoka
Ariki
A
Ochiai
Takehiko
T
Shikama
Keiji
K
D000595:Amino Acid Sequence; D000818:Animals; D002175:Candida / Q000737:chemistry*; D002942:Circular Dichroism; D049308:Dioxygenases; D005656:Fungal Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006420:Hemeproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011487:Protein Conformation; D029701:Saccharomyces cerevisiae Proteins; D013053:Spectrophotometry; D014907:Whales
D005656:Fungal Proteins; D006420:Hemeproteins; D029701:Saccharomyces cerevisiae Proteins; D049308:Dioxygenases; C101796:YHB1 protein, S cerevisiae