Inhibitors of difficult protein-protein interactions identified by high-throughput screening of multiprotein complexes.
Inhibitors of difficult protein-protein interactions identified by high-throughput screening of multiprotein complexes.
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DOI：
10.1021/cb400356m
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发表时间：
2013-09-20
影响因子：
4
通讯作者：
Gestwicki JE
中科院分区：
文献类型：
--
作者：
Cesa LC;Patury S;Komiyama T;Ahmad A;Zuiderweg ERP;Gestwicki JE
蛋白质-蛋白质相互作用（PPI）在细胞功能的各个方面都很重要，并且有兴趣寻找这些接触的抑制剂。然而，具有弱亲和力和/或大界面的PPI传统上对抑制剂的发现更有抵抗力，部分原因是开发允许直接测量这些物理相互作用的高通量筛选（HTS）方法更具挑战性。在这里，我们探讨了弱PPI的功能后果是否可以用作结合的替代品。作为模型，我们使用细菌ATP酶DnaK及其伙伴DnaJ和GrpE。DnaJ和GrpE都结合DnaK并催化加速其ATP循环，因此我们使用刺激的核苷酸周转来间接报道这些PPI。在试点筛选中，我们确定了阻断DnaJ或GrpE激活DnaK的化合物。有趣的是，这些分子中至少有一种阻断了DnaK与DnaJ的结合，而另一种化合物破坏了DnaK和GrpE之间的变构而不改变物理相互作用。这些发现表明，在某些情况下，重构的多蛋白复合物的活性可用于鉴定挑战性PPI的变构抑制剂。
Protein-protein interactions (PPIs) are important in all aspects of cellular function and there is interest in finding inhibitors of these contacts. However, PPIs with weak affinities and/or large interfaces have traditionally been more resistant to the discovery of inhibitors, partly because it is more challenging to develop high throughput screening (HTS) methods that permit direct measurements of these physical interactions. Here, we explored whether the functional consequences of a weak PPI might be used as a surrogate for binding. As a model, we used the bacterial ATPase DnaK and its partners DnaJ and GrpE. Both DnaJ and GrpE bind DnaK and catalytically accelerate its ATP cycling, so we used stimulated nucleotide turnover to indirectly report on these PPIs. In pilot screens, we identified compounds that block activation of DnaK by either DnaJ or GrpE. Interestingly, at least one of these molecules blocked binding of DnaK to DnaJ, while another compound disrupted allostery between DnaK and GrpE without altering the physical interaction. These findings suggest that the activity of a reconstituted multi-protein complex might be used in some cases to identify allosteric inhibitors of challenging PPIs.
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影响因子：
3
作者：
Makley LN;Gestwicki JE
通讯作者：
Gestwicki JE
影响因子：
2.9
作者：
Packschies, L;Theyssen, H;Reinstein, J
通讯作者：
Reinstein, J
影响因子：
8
作者：
Mayer, MP;Bukau, B
通讯作者：
Bukau, B
DOI：
10.1126/science.1198701
发表时间：
2011-05-06
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Good MC;Zalatan JG;Lim WA
通讯作者：
Lim WA
影响因子：
56.9
作者：
Harrison, CJ;HayerHartl, M;Kuriyan, J
通讯作者：
Kuriyan, J