Substrate deconstruction and the nonadditivity of enzyme recognition.
Substrate deconstruction and the nonadditivity of enzyme recognition.
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DOI：
10.1021/ja501354q
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发表时间：
2014-05-21
影响因子：
15
通讯作者：
Shoichet BK
中科院分区：
文献类型：
--
作者：
Barelier S;Cummings JA;Rauwerdink AM;Hitchcock DS;Farelli JD;Almo SC;Raushel FM;Allen KN;Shoichet BK
预测未知功能的酶的底物是一个主要的后基因组挑战。底物的发现，抑制剂的发现，是由我们的能力，探索化学型的限制;它会扩大的数量级，如果反应位点可以用片段探测，而不是完全精心制作的基板，是抑制剂的发现。为了探索这种方法的可行性，将来自三个不同超家族的六种酶的底物解构成41个重叠片段，测试其活性或结合。令人惊讶的是，即使是那些含有关键反应基团的片段也几乎没有活性，并且大多数片段在捕获大多数底物特征之前都没有可测量的结合。从被识别的底物上去除一个原子通常会使催化识别减少6个对数级。为了探索在原子分辨率下的识别，通过X射线晶体学确定β-内酰胺酶底物头孢噻吩的三个片段复合物的结构。底物的发现可能很难降低到片段水平，这对功能发现和酶对代谢物混杂的耐受性有影响。实际上，这项研究支持开发的图书馆充分阐述的代谢产物作为探针的酶功能，目前还不存在。
Predicting substrates for enzymes of unknown function is a major postgenomic challenge. Substrate discovery, like inhibitor discovery, is constrained by our ability to explore chemotypes; it would be expanded by orders of magnitude if reactive sites could be probed with fragments rather than fully elaborated substrates, as is done for inhibitor discovery. To explore the feasibility of this approach, substrates of six enzymes from three different superfamilies were deconstructed into 41 overlapping fragments that were tested for activity or binding. Surprisingly, even those fragments containing the key reactive group had little activity, and most fragments did not bind measurably, until they captured most of the substrate features. Removing a single atom from a recognized substrate could often reduce catalytic recognition by 6 log-orders. To explore recognition at atomic resolution, the structures of three fragment complexes of the β-lactamase substrate cephalothin were determined by X-ray crystallography. Substrate discovery may be difficult to reduce to the fragment level, with implications for function discovery and for the tolerance of enzymes to metabolite promiscuity. Pragmatically, this study supports the development of libraries of fully elaborated metabolites as probes for enzyme function, which currently do not exist.
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影响因子：
7.3
作者：
Friberg, Anders;Vigil, Dominico;Zhao, Bin;Daniels, R. Nathan;Burke, Jason P.;Garcia-Barrantes, Pedro M.;Camper, DeMarco;Chauder, Brian A.;Lee, Taekyu;Olejniczak, Edward T.;Fesik, Stephen W.
通讯作者：
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影响因子：
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通讯作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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影响因子：
64.8
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