Phosphate activation in the ground state of purine nucleoside phosphorylase
Phosphate activation in the ground state of purine nucleoside phosphorylase
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DOI：
10.1021/ja0570281
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发表时间：
2006-06-21
影响因子：
15
通讯作者：
Schramm, Vern L.
中科院分区：
文献类型：
--
作者：
Deng, Hua;Murkin, Andrew S.;Schramm, Vern L.
本文用傅里叶变换红外光谱法研究了人嘌呤核苷磷酸化酶（PNP）上的磷酸和核糖1-磷酸（R1 P），并与过渡态类似物上的磷酸进行了比较。结合的磷酸盐是双阴离子的，但以两种不同的结合模式存在，具有相似的结合亲和力。结合R1 P的磷酸盐也是双阴离子。结合的R1 P即使在没有核碱基的情况下也缓慢水解为核糖和磷酸。结合的R1 P中的C-OP键被断裂，与PNP催化的反应相同。游离R1 P经历C-OP和CO-P溶剂分解。在两个PNP（.）R1 P复合物和（PNPPO 4）-P-的一种形式。复杂. PNP中心点R1 P复合物中P-O键的平均氢键强度小于在水中的，但大于PNP中心点PO 4复合物。结合R1 P的水解可以通过结合R1 P中的磷酸部分的变形来引发。结合R1 P的磷酸部分上的不利相互作用通过R1 P从PNP解离或通过水解为核糖和磷酸而减轻。PNP中心点PO 4复合物中的两种形式的结合磷酸被解释为定位为核苷合成方向上的产物和磷酸解反应中的反应物的磷酸;它们的相互转化可以通过质子从一个PO键转移到另一个PO键来发生。与过渡态类似物结合的磷酸盐的电子结构与米氏络合物中的电子结构有很大不同。
Phosphate and ribose 1-phosphate (R1P) bound to human purine nucleoside phosphorylase (PNP) have been studied by FTIR spectroscopy for comparison with phosphate bound with a transition state analogue. Bound phosphate is dianionic but exists in two distinct binding modes with similar binding affinities. The phosphate of bound R1P is also dianionic. Bound R1P slowly hydrolyzes to ribose and phosphate even in the absence of nucleobase. The C-OP bond is cleaved in bound R1P, the same as in the PNP-catalyzed reaction. Free R1P undergoes both C-OP and CO-P solvolysis. A hydrogen bond to one P - O group is stronger than those to the other two P - O groups in both the PNP(.)R1P complex and in one form of the (PNPPO4)-P-. complex. The average hydrogen bond strength to the P - O bonds in the PNP center dot R1P complex is less than that in water but stronger than that in the PNP center dot PO4 complex. Hydrolysis of bound R1P may be initiated by distortion of the phosphate moiety in bound R1P. The unfavorable interactions on the phosphate moiety of bound R1P are relieved by dissociation of R1P from PNP or by hydrolysis to ribose and phosphate. The two forms of bound phosphate in the PNP center dot PO4 complex are interpreted to be phosphate positioned as the product in the nucleoside synthesis direction and as the reactant in the phosphorolysis reaction; their interconversion can occur by the transfer of a proton from one PO bond to another. The electronic structure of phosphate bound with a transition state analogue differs substantially from that in the Michaelis complexes.