Centeno J A
Department of Environmental and Toxicologic Pathology, Armed Forces Institute of Pathology, Washington, DC 20306-6000.
Arch Biochem Biophys. 1992 Feb 1;292(2):624-8. doi: 10.1016/0003-9861(92)90041-t.
The resonance Raman spectra of deoxygenated solutions of mixed-valence cyanide-bound and fully reduced cytochrome oxidase derivatives that have been reduced in the presence of aqueous or solid sodium dithionite exhibit two new low-frequency lines centered at 474 and 590 cm-1. These lines were not observed when the reductant system was changed to a solution containing ascorbate and N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD). Under enzyme turnover conditions, the addition of dithionite to the reoxidized protein (the 428-nm or "oxygenated" form) increases the intensity of these lines, while reoxidation and rereduction of the enzyme in the presence of ascorbate/TMPD resulted in the absence of both lines. Our data suggest that both lines must have contributions from species formed from aqueous dithionite, presumably the SO2 species, since these two lines are also observed in the Raman spectrum of a solution of aqueous dithionite, but not in the spectrum of an ascorbate/TMPD solution. Since heme metal-ligand stretch vibrations are expected to appear in the low-frequency region from 215 to 670 cm-1, our results indicate that special care should be exercised during the interpretation of the cytochrome a3 resonance Raman spectrum.
在水合或固态连二亚硫酸钠存在下还原得到的混合价态氰化物结合型和完全还原型细胞色素氧化酶衍生物的脱氧溶液的共振拉曼光谱显示出两条新的低频谱线，中心位于474和590 cm-1处。当将还原剂体系换成含有抗坏血酸和N,N,N',N'-四甲基对苯二胺（TMPD）的溶液时，未观察到这些谱线。在酶周转条件下，向再氧化的蛋白质（428 nm或“氧化态”形式）中添加连二亚硫酸钠会增加这些谱线的强度，而在抗坏血酸/TMPD存在下对酶进行再氧化和再还原则导致这两条谱线均不存在。我们的数据表明，这两条谱线必定都来自水合连二亚硫酸钠形成的物种的贡献，大概是SO2物种，因为在连二亚硫酸钠水溶液的拉曼光谱中也观察到了这两条谱线，但在抗坏血酸/TMPD溶液的光谱中未观察到。由于血红素金属-配体伸缩振动预计出现在215至670 cm-1的低频区域，我们的结果表明，在解释细胞色素a3共振拉曼光谱时应格外小心。
