Confirmation of a Protein-Protein Interaction in the Pantothenate Biosynthetic Pathway by Using Sortase-Mediated Labelling.
Confirmation of a Protein-Protein Interaction in the Pantothenate Biosynthetic Pathway by Using Sortase-Mediated Labelling.
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DOI：
10.1002/cbic.201500547
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发表时间：
2016-04-15
期刊：
影响因子：
--
通讯作者：
Webb ME
中科院分区：
文献类型：
--
作者：
Morrison PM;Balmforth MR;Ness SW;Williamson DJ;Rugen MD;Turnbull WB;Webb ME
High-throughput studies have been widely used to identify protein-protein interactions however the veracity of few of these candidate interactions have been demonstrated in vitro. We use a combination of isothermal titration calorimetry and fluorescence anisotropy to screen candidate interactions within the pantothenate biosynthetic pathway. In particular, we observe no interaction between the subsequent enzyme in the pathway, pantothenate synthetase (PS) and aspartate decarboxylase but do observe interaction of PS and the putative Nudix hydrolase, YfcD. Confirmation of the interaction by fluorescence anisotropy was dependent upon labelling of an adventitiously formed glycine on the protein N-terminal affinity purification tag using Sortase. Subsequent formation of the protein-protein complex led to apparent restriction of the dynamics of this tag, suggesting that this approach could be generally applied to a subset of other protein-protein interaction complexes.
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DOI：
10.1016/j.bbrc.2012.08.084
发表时间：
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影响因子：
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