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- W2056839429 abstract "Escherichia coli phosphofructokinase, a homotetrameric enzyme with a single tryptophan (W311) per subunit, experiences allosteric inhibition by phosphoenolpyruvate (PEP). The four active sites and four allosteric sites beget four unique heterotropic allosteric interactions, identified by the distance between the two sites; 23A, 30A, 33A and 45A, respectively. Each of the four interactions can be isolated through formation of hybrids containing three inactive tryptophan-minus subunits, and one active subunit retaining a single tryptophan that acts as a fluorescence reporter. Intrinsic fluorescence anisotropy and lifetime measurements are used to calculate the rotational correlation time of various tryptophan-shift mutants, which is used to evaluate how PEP inhibition perturbs the local dynamics around the tryptophan at specific locations. The purpose of this study is to compare the 23A and 30A interaction in terms of the ligand-induced perturbations of local dynamics throughout the enzyme. Rotational correlation times have been determined for W233 in unligated enzyme, enzyme with fructose 6-phosphate (F6P) or PEP bound, and enzyme with both F6P and PEP bound, in both the 23A and 30A interaction. In the 30A interaction, perturbations to the rotational correlation time of W233 with both ligands bound (−8.2±0.4ns) is not a sum of the effects from the individual binding of F6P (8.4±0.4) and PEP (−2.4±0.4) alone. This observation suggests that the region around W233 is involved in the allosteric coupling between F6P and PEP. The rotational correlation time of W233 is relatively undisturbed by F6P, PEP or both binding in the 23A interaction (−1.3±0.1, 0.5±.2, 0.6±0.1ns respectively). These data suggest that the region around W233 is important for transmission of the allosteric signal in the 30A interaction but not in the 23A interaction. Funding: NIH-GM33216, and Welch Foundation A1543." @default.
- W2056839429 created "2016-06-24" @default.
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- W2056839429 date "2012-01-01" @default.
- W2056839429 modified "2023-09-26" @default.
- W2056839429 title "Ligand Induced Dynamic Perturbations Highlight Important Regions Leading to Inhibition of Escherichia Coli Phosphofructokinase by Phosphoenolpyruvate" @default.
- W2056839429 doi "https://doi.org/10.1016/j.bpj.2011.11.377" @default.
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