Enzymology with a Spin-Labeled Phospholipase C: Soluble Substrate Binding by 31P NMR from 0.005 to 11.7 T
Enzymology with a Spin-Labeled Phospholipase C: Soluble Substrate Binding by 31P NMR from 0.005 to 11.7 T
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DOI：
10.1021/bi901190j
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发表时间：
2009-09-08
期刊：
影响因子：
2.9
通讯作者：
Roberts, Mary F.
中科院分区：
文献类型：
--
作者：
Pu, Mingming;Feng, Jianwen;Roberts, Mary F.
从0.005到11.7皮重的P-31 NMR弛豫研究用于监测水溶性肌醇1，2-（环）磷酸（cIP）与磷脂酰肌醇特异性磷脂酶C（H82 C，该酶活性位点附近的位置）的自旋标记结合，并确定激活磷脂酰胆碱（PC）分子如何影响这种相互作用。我们表明，在没有界面的情况下，cIP与蛋白质的结合不是限速的，并且与胶束相反，PC囊泡的较低活化可能是由于产物释放受阻。该方法是通用的，可用于确定其他弱结合小分子配体-蛋白质相互作用的距离。
P-31 NMR relaxation studies from 0.005 to 11.7 Tare used to monitor water-soluble inositol 1,2-(cyclic) phosphate (cIP) binding to phosphatidylinositol-specific phospholipase C spin-labeled at H82C, a position near the active site of the enzyme, and to determine how activating phosphatidylcholine (PC) molecules affect this interaction. We show that, in the absence of an interface, cIP binding to the protein is not rate-limiting, and that lower activation by PC vesicles as opposed to micelles is likely due to hindered product release. The methodology is general and could be used for determining distances in other weakly binding small molecule ligand-protein interactions.