Section 42
Chapter 41,249

Resonance Raman study on mutant cytochrome P-450 obtained by site-directed mutagenesis

Egawa, T.; Imai, Y.; Ogura, T.; Kitagawa, T.

Biochimica et Biophysica Acta 1040(2): 211-216


ISSN/ISBN: 0006-3002
PMID: 2119227
DOI: 10.1016/0167-4838(90)90078-t
Accession: 041248738

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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(omega-1)-hydroxylase] (P-450(omega-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 Fe-CO stretching mode (vFe-CO) 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 vFe-CO 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.

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