BINDING OF THE COMPETITIVE INHIBITOR DCDP TO RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE FROM ESCHERICHIA-COLI STUDIED BY H-1-NMR - DIFFERENT PROPERTIES OF THE LARGE PROTEIN SUBUNIT AND THE HOLOENZYME
BINDING OF THE COMPETITIVE INHIBITOR DCDP TO RIBONUCLEOSIDE-DIPHOSPHATE REDUCTASE FROM ESCHERICHIA-COLI STUDIED BY H-1-NMR - DIFFERENT PROPERTIES OF THE LARGE PROTEIN SUBUNIT AND THE HOLOENZYME
复制标题
DOI：
10.1111/j.1432-1033.1992.tb17229.x
复制
发表时间：
1992-09-15
期刊：
影响因子：
--
通讯作者：
EHRENBERG, A
中科院分区：
文献类型：
--
作者：
ALLARD, P;KUPRIN, S;EHRENBERG, A
来自大肠杆菌的核糖核苷二磷酸还原酶（ec1.17.4.1）由两个不相同的亚基，蛋白R1和R2组成。利用h -1核磁共振光谱研究了产物dCDP与蛋白R1和全酶R1R2的结合。在25℃的温度下，效应物dTTP存在时，发现dCDP在结合位点和溶剂之间快速交换，导致dCDP共振变宽。当蛋白R1和R2同时存在时，复合物R1R2形成，观察到的增宽比单独存在蛋白R1时要小。当[R2]/[R1]比值超过1时，线宽未进一步减小。的结合常数dCDP R1或R1R2是相同的,K (d) = 0.9毫米。小扩大dCDP共振观察到的复杂R1R2与R1相比可能解释的组合两个效果:(a)蛋白质的总体下跌时间将增加从R1 R1R2时,这将导致扩大相应增加,和(b)两方面的结合位点的数量快速减少交换,这将减少扩大0.5倍。与天然R2相比，不含铁的R2 （apoR2）的作用降低了，可能是由于一些变性蛋白，而R2的c端肽根本没有引起任何狭窄。
Ribonucleoside-diphosphate reductase (EC 1.17.4.1) from Escherichia coli consists of two nonidentical subunits, proteins R1 and R2. The binding of the product dCDP to protein R1 and to the holoenzyme R1R2 has been studied by means of H-1-NMR spectroscopy. In presence of the effector dTTP at 25-degrees-C, dCDP was found to be in rapid exchange between the binding sites and the solvent which results in a broadening of the dCDP resonances. When both proteins R1 and R2 are present, so that the complex R1R2 is formed, a smaller broadening is observed than with protein R1 alone. No further linewidth decrease was observed when the [R2]/[R1] ratio exceeded 1. The binding constant of dCDP to R1 or R1R2 is the same, K(d) = 0.9 mM. The smaller broadening of the dCDP resonances observed with the complex R1R2 as compared with R1 may be explained by the combination of two effects: (a) the overall tumbling time of the protein will increase when going from R1 to R1R2, which will cause the broadening to increase correspondingly, and (b) a twofold decrease of the number of binding sites in rapid exchange, which will decrease the broadening by a factor of 0.5. The effect of R2 without iron (apoR2) is reduced compared with native R2, probably because of some denatured proteins, while a C-terminal peptide from R2 did not cause any narrowing at all.