Othman S, Le Lirzin A, Desbois A
Département de Biologie Cellulaire et Moléculaire, Centre d'Etudes de Saclay, Gif-sur-Yvette, France.
Biochemistry. 1993 Sep 21;32(37):9781-91. doi: 10.1021/bi00088a033.
The visible absorption and Soret-excited resonance Raman spectra of ferrous microperoxidase-8 [MP8(II)], an octapeptide containing a heme c, are reported. These spectroscopies indicate that MP8(II), dissolved in aqueous buffered solutions, forms low-spin six-coordinated complexes in the 7-14 pH range. Intermolecular bonding interactions of MP8(II) in water account for this behavior. On the contrary, when the hemopeptide is dispersed in aqueous solutions containing detergent or an alcohol, the spectroscopic data show that the iron atom of MP8(II) is essentially high-spin five-coordinated in accordance with a monomeric structure of MP8(II). In addition to a high-spin signature to the heme skeletal modes, the high-frequency regions of resonance Raman spectra characterize an electronic influence of the thioether bridges on the frequency of stretching modes of C beta-C beta bonds (nu 2, nu 11, and nu 29). On the other hand, the low-frequency Raman spectra of monomeric MP8(II) at pH 7.5 present significant differences in the 150-250-cm-1 regions depending upon the solvent composition (pH, presence or absence of detergent, alcohol). These effects are attributed to frequency variations of the Fe-N(His)-involving mode which indicate changes in the H-bonding interactions of the axial His and therefore solvent-dependent changes of the octapeptide conformation. Our resonance Raman data further show that the axial His of monomeric MP8(II) could be totally deprotonated in aqueous cetyltrimethylammonium bromide solution at very alkaline pH (pKa = 13.3). The vibrational data (100-1700 cm-1) obtained for the various monomeric forms of MP8(II) are expected to be useful for determining the heme structure and environment in reduced c'-type cytochromes. Comparisons of resonance Raman data with X-ray crystallographic data available for different hemoproteins allow us to evaluate the ionization and H-bonding states of the His bound to the high-spin five-coordinated hemes. These data are discussed in terms of proximal influence of protein-His-heme interactions on the determination and the regulation of a particular biological function.
报道了含血红素c的八肽亚铁微过氧化物酶-8 [MP8(II)]的可见吸收光谱和Soret激发共振拉曼光谱。这些光谱表明，溶解在水性缓冲溶液中的MP8(II)在7 - 14的pH范围内形成低自旋六配位络合物。MP8(II)在水中的分子间键合相互作用解释了这种行为。相反，当血红肽分散在含有去污剂或醇的水溶液中时，光谱数据表明，根据MP8(II)的单体结构，MP8(II)的铁原子基本上是高自旋五配位的。除了血红素骨架模式的高自旋特征外，共振拉曼光谱的高频区域还表征了硫醚桥对Cβ - Cβ键拉伸模式频率（ν2、ν11和ν29）的电子影响。另一方面，在pH 7.5时，单体MP8(II)的低频拉曼光谱在150 - 250 cm-1区域根据溶剂组成（pH、去污剂的存在与否、醇）呈现出显著差异。这些效应归因于涉及Fe-N(His)模式的频率变化，这表明轴向His的氢键相互作用发生了变化，因此八肽构象也发生了溶剂依赖性变化。我们的共振拉曼数据进一步表明，在非常碱性的pH（pKa = 13.3）下，单体MP8(II)的轴向His在十六烷基三甲基溴化铵水溶液中可能完全去质子化。为MP8(II)的各种单体形式获得的振动数据（100 - 1700 cm-1）有望用于确定还原型c'-型细胞色素中的血红素结构和环境。将共振拉曼数据与不同血红蛋白质的X射线晶体学数据进行比较，使我们能够评估与高自旋五配位血红素结合的His的电离和氢键状态。将根据蛋白质-His-血红素相互作用对特定生物学功能的确定和调节的近端影响来讨论这些数据。
