Elucidation of the Differences between the 430- and 455-nm Absorbing Forms of P450-Isocyanide Adducts by Resonance Raman Spectroscopy

Takeshi Tomita, Seiji Ogo, Tsuyoshi Egawa, Hideo Shimada, Noriaki Okamoto, Yoshio Imai, Yoshihito Watanabe, Yuzuru Ishimura, Teizo Kitagawa
2001 Journal of Biological Chemistry  
Alkylisocyanide adducts of microsomal P450 exist in two interconvertible forms, each giving the Soret maximum around 430 or 455 nm. This is demonstrated with a rabbit liver P450 2B4. Resonance Raman spectra of the 430-and 455-nm forms were examined for typical P450s of the two types as well as for P450 2B4 because the 430-nm form of P450 2B4 is liable to change into P420. P450cam and P450nor were selected as a model of the 430-and 455-nm forms, respectively. For the n-butyl isocyanide (CNBu)
more » ... uct, the Fe(II)-CNBu stretching band was observed for the first time at 480/467 cm ؊1 for P450cam and at 471/459 cm ؊1 for P450nor with their 12 CNBu/ 13 CNBu derivatives. For P450cam, but not P450nor, other 13 C isotope-sensitive bands were observed at 412/402, 844/835, and 940/926 cm ؊1 . The C-N stretching mode was identified by Fourier transform IR spectroscopy at 2116/2080 cm ؊1 for P450cam and at 2148/ 2108 cm ؊1 for P450nor for the 12 C/ 13 C derivatives. These findings suggest that the binding geometry of isocyanide differs between the two forms-bent and linear structures for P450cam-CNBu and P450nor-CNBu, respectively. In contrast, in the ferric state, the Raman 13 C isotopic frequency shifts, and the IR C-N stretching frequencies (2213/2170 and 2215/2172 cm ؊1 ) were similar between P450cam and P450nor, suggesting similar bent structures for both.
doi:10.1074/jbc.m104932200 pmid:11459844 fatcat:eqjigvossfb4ppstggnvj347am