The Relationship between Enzyme Conformational Change, Proton Transfer, and Phosphoryl Transfer in ß-Phosphoglucomutase
The Relationship between Enzyme Conformational Change, Proton Transfer, and Phosphoryl Transfer in ß-Phosphoglucomutase
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α-磷酸葡萄糖变位酶中酶构象变化、质子转移和磷酰基转移之间的关系
DOI：
10.1021/acscatal.1c01389
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发表时间：
2021
期刊：
影响因子：
12.9
通讯作者：
Robertson A
中科院分区：
文献类型：
--
作者：
Robertson A
磷酰基转移反应中质子转移的时间和作用的分子细节知之甚少。在这里，我们结合了QM模型，实验NMR测量和X射线结构，以建立一个原型磷酰基转移酶，βPGM，从一个非常封闭的近攻击构象到一个完全封闭的过渡态类似物（TSA）构象触发了两个部分质子转移从一般的酸-碱残基转化为离去基团氧，转移的磷酰基从离去基团氧部分解离。质子转移继续进行，但在TSA构象的酶的磷酰基转移的整个反应路径中没有完成。此外，使用相互作用量子原子（IQA）和相对能量梯度（REG）分析方法，我们观察到，质子的位置的变化和相应的质子和磷原子之间的静电斥力增加提供了一个刺激磷酰基转移串联在一起的离去基团氧原子上的负电荷密度的减少。β PGMWT和β PGMD 10 NTSA复合物的溶液相19 F NMR测量结果与等效QM模型之间的一致性证实了两种变体中G6 P的质子化状态，验证了所采用的QM模型。此外，使用高分辨率X射线晶体结构证实了QM模型对AlF 4响应于质子位置的扭曲的预测，不仅为QM模型提供了额外的验证，而且还进一步建立了金属氟化物作为活性位电荷密度分布的高灵敏度实验预测因子。
Molecular details for the timing and role of proton transfer in phosphoryl transfer reactions are poorly understood. Here, we have combined QM models, experimental NMR measurements, and X-ray structures to establish that the transition of an archetypal phosphoryl transfer enzyme, βPGM, from a very closed near-attack conformation to a fully closed transition state analogue (TSA) conformation triggers both partial proton transfer from the general acid–base residue to the leaving group oxygen and partial dissociation of the transferring phosphoryl group from the leaving group oxygen. Proton transfer continues but is not completed throughout the reaction path of the phosphoryl transfer with the enzyme in the TSA conformation. Moreover, using interacting quantum atoms (IQA) and relative energy gradient (REG) analysis approaches, we observed that the change in the position of the proton and the corresponding increased electrostatic repulsion between the proton and the phosphorus atom provide a stimulus for phosphoryl transfer in tandem with a reduction in the negative charge density on the leaving group oxygen atom. The agreement between solution-phase19F NMR measurements and equivalent QM models of βPGMWTand βPGMD10NTSA complexes confirms the protonation state of G6P in the two variants, validating the employed QM models. Furthermore, QM model predictions of an AlF4distortion in response to the proton position are confirmed using high resolution X-ray crystal structures, not only providing additional validation to the QM models but also further establishing metal fluorides as highly sensitive experimental predictors of active-site charge density distributions.