La Mar G N, Krishnamoorthi R
Biophys J. 1983 Nov;44(2):177-83. doi: 10.1016/S0006-3495(83)84289-1.
Four exchangeable protons with large hyperfine shifts are assigned in the heme pocket of sperm whale met-cyano myoglobin reconstituted with heme possessing acetyl groups, ethyl groups, bromines, and hydrogens at the 2,4 position, using both relaxation and chemical-shift data. The four protons arise from the ring NH's of the proximal (F8), distal (E7), and FG2 histidines, and the peptide NH of His F8. The similarity of all chemical shifts to those of the native protein as well as the invariance of the relaxation rates of the distal histidyl ring NH dictate essentially the same structure for the heme cavity of both native and reconstituted proteins. The exchange rates with bulk water of the four labile proteins in each modified protein were determined by saturation-transfer and line width methods. All four labile protons were found to have the same exchange rate as in the native protein for acetyl and ethyl 2,4 substituents; the two resolved labile protons in the derivative with 2,4 bromine were also unchanged. The reconstituted protein with hydrogens at the 2,4 position exhibited slower exchange rates for three of the four protons, indicating an increased dynamic stability of the heme pocket in the absence of bulky 2,4 substituents.
利用弛豫和化学位移数据，在具有乙酰基、乙基、溴和2,4位氢的血红素重构的抹香鲸高铁氰化肌红蛋白的血红素口袋中，确定了四个具有大超精细位移的可交换质子。这四个质子来自近端（F8）、远端（E7）和FG2组氨酸的环NH以及His F8的肽NH。所有化学位移与天然蛋白质的化学位移相似，以及远端组氨酸环NH弛豫率的不变性，表明天然和重构蛋白质的血红素腔结构基本相同。通过饱和转移和线宽方法测定了每种修饰蛋白质中四个不稳定质子与大量水的交换率。发现对于乙酰基和乙基2,4取代基，所有四个不稳定质子的交换率与天然蛋白质中的相同；在具有2,4溴的衍生物中两个分辨出的不稳定质子也没有变化。在2,4位具有氢的重构蛋白质中，四个质子中的三个表现出较慢的交换率，这表明在没有庞大的2,4取代基的情况下，血红素口袋的动态稳定性增加。
