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A New Approach for the Identification of Allosteric Binding Sites in Proteins
Author(s) -
Striebeck Petra,
Manstein Dietmar J.
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.964.6
Subject(s) - microscale thermophoresis , allosteric regulation , chemistry , metabolite , small molecule , binding site , myosin , biochemistry , biophysics , folding (dsp implementation) , allosteric enzyme , plasma protein binding , enzyme , function (biology) , binding domain , biology , microbiology and biotechnology , electrical engineering , engineering
Recent results indicate that small‐molecule‐protein interactions modulate the function of a large fraction of the enzyme complement in eukaryotic cells. In particular, modulation of enzyme functions by small metabolites that are not their substrates or products is increasingly recognized as a mechanism by which cells can adapt to changes in environmental conditions. While the effect of metabolite‐binding on conformational transitions and thus protein folding, stability, and catalytic activity is clearly of considerable importance, studying metabolite‐protein interactions is in most instances impeded by the low‐affinity nature of the interactions, their moderate effect on enzyme activity, and the complex nature of their interference with substrate turnover. Here, we describe an approach for the efficient mapping of the binding sites of nutrient polyphenols and small dye molecules to the 80 kDa myosin motor domain. We speculate that their binding occurs preferentially with sites that are important for protein‐metabolite interactions and can also be exploited as binding sites for therapeutic drugs. To determine the affinity of the prolific binders for the myosin‐2 motor domain and to select the most suitable compounds, we performed microscale thermophoresis experiments. Subsequently, we determined the structure of the binding pockets for the most promising compounds by X‐ray crystallography.