Pinpointing the l-phenylalanine binding sites of TyrR using biosensors and computer-aided simulation
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To determine the binding sites for l-phenylalanine in TyrR protein via a rational mutation analysis combining biosensors and computer-aided simulation.
TyrR protein of Escherichia coli is the chief transcriptional regulator of several genes essential for the biosynthesis and transport of aromatic amino acids. The identification of ligand-binding sites is often the starting point for protein function annotation and structure-based protein design. Here we combined computer-aided prediction methods and biosensors to identify the ligand-binding sites for l-Phe in TyrR protein.
Residues at positions 160, 173 and 184 of TyrR protein are important for transcriptional activation of target genes tyrP induced by l-Phe, which indicates that they are the bona fide l-Phe binding sites of TyrR protein.
KeywordsTyrR protein l-Phe binding site Biosensor Computer-aided simulation
This work was financially supported by National Key R&D Program of China (2018YFD0901001), Tianjin Science Fund for Distinguished Young Scholars (17JCJQJC45300), Nature Science Foundation of Tianjin City (CN) (16JCYBJC23500), Tianjin Science and Technology Project (15PTCYSY00020) and the Science and Technology Service Network (STS) Initiative of Chinese Academy of Sciences.
Supplementary Table 1—Strains and plasmids used in this study.
Supplementary Table 2—Primers used in this study.
Supplementary Figure 1—The effect of 184 site of TyrR on the binding mode of l-phe. The representative conformation of l-phe with 173 and 160 sites is verified by mutation S184L/V. The mutations S184L and S184V would clash with the benzene ring of l-Phe when binds 173 and 160 sites.
Supplementary Figure 2—Representative conformations of three clusters in MD simulations for 15 site of TyrR protein.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
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