Nagatomo Shigenori, Saito Kazuya, Yamamoto Kohji, Ogura Takashi, Kitagawa Teizo, Nagai Masako
Department of Chemistry, Faculty of Pure and Applied Sciences, University of Tsukuba , Tsukuba, Ibaraki 305-8571, Japan.
Research Center for Development of Far-Infrared Region, University of Fukui , Fukui, Fukui 910-8507, Japan.
Biochemistry. 2017 Nov 21;56(46):6125-6136. doi: 10.1021/acs.biochem.7b00733. Epub 2017 Nov 7.
Following a previous detailed investigation of the β subunit of αβ human adult hemoglobin (Hb A), this study focuses on the α subunit by using three natural valency hybrid α(Fe-deoxy/O)β(Fe) hemoglobin M (Hb M) in which O cannot bind to the β subunit: Hb M Hyde Park (β92His → Tyr), Hb M Saskatoon (β63His → Tyr), and Hb M Milwaukee (β67Val → Glu). In contrast with the β subunit that exhibited a clear correlation between O affinity and Fe-His stretching frequencies, the Fe-His stretching mode of the α subunit gave two Raman bands only in the T quaternary structure. This means the presence of two tertiary structures in α subunits of the αβ tetramer with T structure, and the two structures seemed to be nondynamical as judged from terahertz absorption spectra in the 5-30 cm region of Hb M Milwaukee, α(Fe-deoxy)β(Fe). This kind of heterogeneity of α subunits was noticed in the reported spectra of a metal hybrid Hb A like α(Fe-deoxy)β(Co) and, therefore, seems to be universal among α subunits of Hb A. Unexpectedly, the two Fe-His frequencies were hardly changed with a large alteration of O affinity by pH change, suggesting no correlation of frequency with O affinity for the α subunit. Instead, a new Fe-His band corresponding to the R quaternary structure appeared at a higher frequency and was intensified as the O affinity increased. The high-frequency counterpart was also observed for a partially O-bound form, α(Fe-deoxy)α(Fe-O)β(Fe)β(Fe), of the present Hb M, consistent with our previous finding that binding of O to one α subunit of T structure αβ tetramer changes the other α subunit to the R structure.
在先前对成人αβ型人血红蛋白（Hb A）β亚基进行详细研究之后，本研究聚焦于α亚基，使用了三种天然价态杂合的α(Fe-脱氧/O)β(Fe)血红蛋白M（Hb M），其中O无法与β亚基结合：海德公园血红蛋白M（β92位组氨酸→酪氨酸）、萨斯卡通血红蛋白M（β63位组氨酸→酪氨酸）和密尔沃基血红蛋白M（β67位缬氨酸→谷氨酸）。与β亚基在O亲和力和Fe-组氨酸伸缩频率之间呈现明显相关性不同，α亚基的Fe-组氨酸伸缩模式仅在T四级结构中给出两条拉曼带。这意味着具有T结构的αβ四聚体的α亚基中存在两种三级结构，并且从密尔沃基血红蛋白M（α(Fe-脱氧)β(Fe)）在5 - 30 cm区域的太赫兹吸收光谱判断，这两种结构似乎是非动态的。在报道的如α(Fe-脱氧)β(Co)这种金属杂合Hb A的光谱中也注意到了α亚基的这种异质性，因此，这似乎在Hb A的α亚基中普遍存在。出乎意料的是，随着pH变化导致O亲和力大幅改变时，这两个Fe-组氨酸频率几乎没有变化，表明频率与α亚基的O亲和力无关。相反，对应于R四级结构的一条新的Fe-组氨酸带在更高频率出现，并且随着O亲和力增加而增强。在本Hb M的部分O结合形式α(Fe-脱氧)α(Fe-O)β(Fe)β(Fe)中也观察到了高频对应物，这与我们之前的发现一致，即O与T结构αβ四聚体的一个α亚基结合会使另一个α亚基转变为R结构。
