For the first time, Fourier transform infrared spectroscopy has been applied to cytochrome P-450 to analyze the protein secondary structure. From Fourier self-deconvolution and fitting the infrared spectra in the amide I' region (1600-1700 cm-1), we estimate 44% alpha-helix, 31% beta-sheet, and 18% turns for substrate-free cytochrome P-450cam. In the presence of camphor, 54% alpha-helix and 310-helix, 21% beta-sheet, and 21% turns are obtained which agree with the crystallographic data of 53% alpha-helix, 19% beta-sheet, and 16% turns [Poulos, T. L., Finzel, B. C., & Howard, A. J. (1987) J. Mol. Biol. 195, 687-700]. Cytochrome P-420cam is produced from substrate-free cytochrome P-450cam in two ways(i) by temperature elevation up to 60 degrees C and (ii) by exposure to KSCN up to 1.5 M. The secondary structure composition is determined for each temperature and KSCN concentration and compared with the changes observed in the iron ligand CO stretch vibration bands appearing between 1900 and 2000 cm-1. Thermally induced cytochrome P-420 has an alpha-helix content of 19%, a beta-sheet content of 53%, 14% turns, and 5% antiparallel beta-sheets from intermolecular hydrogen bonds within protein aggregates. The formation of cytochrome P-420 as a function of the KSCN concentration indicates two types of cytochrome P-420. Up to 1 M KSCN, the induced cytochrome P-420 displays only little modification of the secondary structure, whereas at 1.5 M KSCN, larger changes are observed, resulting in 85% cytochrome P-420 without protein precipitation and containing 30% alpha-helix, 48% beta-sheet, and 17% turns. Infrared spectra in the iron ligand CO stretch region show several subconformers for cytochrome P-420. During the cytochrome P-420 formation, the CO stretch modes are shifted to higher frequencies by 3-11 cm-1, with a main feature at about 1964 cm-1, compared to those of substrate-free cytochrome P-450cam-CO.

译文

第一次,傅里叶变换红外光谱已应用于细胞色素P-450,以分析蛋白质的二级结构。通过傅立叶自解卷积并拟合酰胺I'区域(1600-1700 cm-1)中的红外光谱,我们估计无底物的细胞色素P-占44%的α-螺旋,31%的β-折叠和18%的匝数。 450cam。在存在樟脑的情况下,获得了54%的α-螺旋和310螺旋,21%的β-折叠和21%的折光,这与53%的α-螺旋,19%的β-折叠和16%的晶体学数据相符[Poulos,TL,Finzel,BC,and Howard,AJ(1987)J. Mol。生物学195,687-700]。由无底物的细胞色素P-450cam产生细胞色素P-420cam的方法有两种(i)通过升高温度达到60摄氏度和(ii)通过暴露于KSCN达到1.5 M.在每个温度下确定二级结构组成和KSCN浓度,并与在1900至2000cm-1之间出现的铁配体CO拉伸振动带中观察到的变化进行比较。热诱导的细胞色素P-420具有19%的α-螺旋含量,53%的β-折叠层含量,14%匝数和5%的蛋白质聚集体中分子间氢键反平行的β-折叠层。细胞色素P-420的形成随KSCN浓度的变化指示出两种类型的细胞色素P-420。高达1 M KSCN时,诱导的细胞色素P-420仅显示出二级结构的修饰,而在1.5 M KSCN时,观察到较大的变化,导致85%的细胞色素P-420没有蛋白质沉淀并且含有30%的α-螺旋,48%的Beta版表和17%的匝数。铁配体CO延伸区的红外光谱显示了细胞色素P-420的多个亚构型。在细胞色素P-420形成过程中,与无底物的细胞色素P-450cam-CO相比,CO拉伸模式向更高的频率移动了3-11 cm-1,主要特征在大约1964 cm-1。

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