TY - JOUR
T1 - Resonance Raman study on mutant cytochrome P-450 obtained by site-directed mutagenesis
AU - Egawa, T.
AU - Imai, Y.
AU - Ogura, T.
AU - Kitagawa, T.
N1 - Funding Information:
This study was supported by a Grant-in-Aid for Scientific Research on Priority Areas (63635005) from the Ministry of Education, Science and Culture. T.E. is indebted to Professor T. Masuda of Tokyo Metropolitan University for encouragement during the course of this study.
PY - 1990/9/3
Y1 - 1990/9/3
N2 - Resonance Raman spectra were observed for the threonine-301 to serine or valine mutant as well as the wild type of rabbit liver microsomal cytochrome P-450 [laurate(ω - 1)-hydroxylase] (P-450(ω - 1), which were prepared through site-directed mutagenesis. The high-spin marker resonance Raman (RR) bands became similarly stronger for all the P-450s examined in the oxidized form upon addition of laurate, and the RR spectra in the higher frequency region of the oxidized, reduced and CO-adduct forms did not distinctly differ among the P-450s examined. Nevertheless, the FeCO stretching mode (νFeCO) of the CO adduct exhibited an upshift for the valine mutant, suggesting positional proximity of Thr-301 to bound CO like Thr-252 of P-450cam, in agreement with the expectation from the sequence analysis. The νFeCO band was shifted to higher frequency upon binding of normal alkyl fatty acids with C10 or longer alkyl chain but little affected by binding of shorter fatty acids.
AB - Resonance Raman spectra were observed for the threonine-301 to serine or valine mutant as well as the wild type of rabbit liver microsomal cytochrome P-450 [laurate(ω - 1)-hydroxylase] (P-450(ω - 1), which were prepared through site-directed mutagenesis. The high-spin marker resonance Raman (RR) bands became similarly stronger for all the P-450s examined in the oxidized form upon addition of laurate, and the RR spectra in the higher frequency region of the oxidized, reduced and CO-adduct forms did not distinctly differ among the P-450s examined. Nevertheless, the FeCO stretching mode (νFeCO) of the CO adduct exhibited an upshift for the valine mutant, suggesting positional proximity of Thr-301 to bound CO like Thr-252 of P-450cam, in agreement with the expectation from the sequence analysis. The νFeCO band was shifted to higher frequency upon binding of normal alkyl fatty acids with C10 or longer alkyl chain but little affected by binding of shorter fatty acids.
KW - Carbonmonoxide adduct
KW - Cytochrome P-450
KW - Laurate(ω - 1)-hydroxylase
KW - Resonance Raman
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U2 - 10.1016/0167-4838(90)90078-T
DO - 10.1016/0167-4838(90)90078-T
M3 - Article
C2 - 2119227
AN - SCOPUS:0025035998
SN - 0167-4838
VL - 1040
SP - 211
EP - 216
JO - Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
JF - Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
IS - 2
ER -