The pKa values of two histidine residues in human haemoglobin, the Bohr effect, and the dipole moments of alpha-helices.
The pKa values of two histidine residues in human haemoglobin, the Bohr effect, and the dipole moments of alpha-helices.
复制标题
人血红蛋白中两个组氨酸残基的 pKa 值、玻尔效应和 α 螺旋的偶极矩。
DOI：
10.1016/0022-2836(85)90016-6
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发表时间：
1985
影响因子：
5.6
通讯作者：
Shih,DT
中科院分区：
文献类型：
--
作者：
Perutz,MF;Gronenborn,AM;Clore,GM;Fogg,JH;Shih,DT
对异常血红蛋白和化学修饰血红蛋白的研究表明，在0. 1 m-NaCl中，人血红蛋白的碱性玻尔效应约有40%是由C-末端组氨酸HC 3（146）β贡献的。在脱氧血红蛋白中，组氨酸的咪唑与天冬氨酸FG 1（94）β形成盐桥，在氧合血红蛋白或碳单氧合血红蛋白中，它接受来自其自身NH基团的氢键。Kilmartinet等人（1973）表明，在0. 2 m-NaCl + 0. 2 m-phosphate中，这种连接的变化使组氨酸的pKa从Hb α的8.0降低到HbCO的7.1，但Russuet al.（1980）声称，在不加NaCl的bis-Tris缓冲液中，HbCO的pKain下降不低于7.85，在这种介质中，C-末端组氨酸对碱性玻尔效应的贡献可以忽略不计。HbCO Cowtown（His HC3（146）β → Leu）、HbCO Wood（His FG4（97）β → Leu）和HbCO Malmostain（His FG4（97）β → Gln）。我们的结果表明Russuetwww.example.com HisHC 3（146）β的共振实际上属于HisFG 4（97）β。al.to虽然在Hb中，HisHC 3（146）β的pK α为8.05 ± 0.05，与离子强度无关，但在HbCO中，其pK α随离子强度的减小而急剧下降，因此在Russuet使用的缓冲液中，pK α为6.2，对碱性玻尔效应的贡献比Kilmartine等人使用的磷酸盐缓冲液大57%。al.it在HbCO A中，His FG 4（97）β对玻尔效应没有贡献，但在His HC 3（146）β已被切割的HbCO中（HbCO des-His），His FG 4（97）β在具有不同pKa值的两种构象之间处于平衡。在HbCO A中，His FG 4（97）β的pKa为7.8，而蛋白质表面游离组氨酸的pKa值约为6.6。这种高pKais是由其与螺旋F和FG的C末端的负极的相互作用引起的。它对应的自由能变化与组氨酸与羧酸根离子相互作用中观察到的自由能变化相同，并证实了α-螺旋的强偶极特性，即使它们位于蛋白质表面也会表现出来。
Studies of abnormal and chemically modified haemoglobins indicate that in 0.1m-NaCl about 40% of the alkaline Bohr effect of human haemoglobin is contributed by the C-terminal histidine HC3(146)β. In deoxyhaemoglobin, the imidazole of this histidine forms a salt bridge with aspartate FG1(94)β, in oxyhaemoglobin or carbonmonoxyhaemoglobin it accepts a hydrogen bond from its own NH group instead. Kilmartinet al.(1973) showed that in 0.2m-NaCl + 0.2m-phosphate this change of ligation lowered the pKaof the histidine from 8.0 in Hb‡to 7.1 in HbCO, but Russuet al.(1980) claimed that in bis-Tris buffer without added NaCl its pKain HbCO dropped no lower than 7.85, and that in this medium the C-terminal histidine made only a negligible contribution to the alkaline Bohr effect.We have compared the histidine resonances of HbCO A with those of three abnormal haemoglobins: HbCO Cowtown (His HC3(146)β → Leu), HbCO Wood (His FG4(97)β → Leu) and HbCO Malmø (His FG4(97)β → Gln). Our results show that the resonance assigned by Russuet al.to His HC3(146)β in fact belongs to His FG4(97)β. Although in Hb the pKaof His HC3(146)β is 8.05 ± 0.05 independent of ionic strength, in HbCO its pKadrops sharply with diminishing ionic strength, so that in the buffer employed by Russuet al.it has a pKaof 6.2 and makes a contribution to the alkaline Bohr effect that is 57%largerthan in the phosphate buffer employed by Kilmartinet al.(1973).In HbCO A, His FG4(97)β does not contribute to the Bohr effect, but in HbCO from which His HC3(146)β has been cleaved (HbCO des-His), His FG4(97)β is in equilibrium between two conformations with different pKavalues. This equilibrium varies with ionic strength and pH, and presumably also with degree of ligation of the haem moiety.In HbCO A, His FG4(97)β has a pKaof 7.8 compared to the pKavalue of about 6.6 characteristic of free histidines at the surface of proteins. This high pKais accounted for by its interaction with the negative pole at the C terminus of helices F and FG. It corresponds to a free energy change of the same order as that observed in the interaction of histidines with carboxylate ions and confirms the strongly dipolar character of α-helices, which manifests itself even when they lie on the surface of the protein.