1. Competing allosteric mechanisms modulate substrate binding in a dimeric enzyme.
- Author
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Freiburger LA, Baettig OM, Sprules T, Berghuis AM, Auclair K, and Mittermaier AK
- Subjects
- Acetyltransferases chemistry, Allosteric Regulation, Calorimetry, Circular Dichroism, Enterococcus faecium chemistry, Models, Molecular, Nuclear Magnetic Resonance, Biomolecular, Protein Binding, Protein Conformation, Protein Multimerization, Substrate Specificity, Thermodynamics, Acetyl Coenzyme A metabolism, Acetyltransferases metabolism, Enterococcus faecium enzymology
- Abstract
Allostery has been studied for many decades, yet it remains challenging to determine experimentally how it occurs at a molecular level. We have developed an approach combining isothermal titration calorimetry, circular dichroism and nuclear magnetic resonance spectroscopy to quantify allostery in terms of protein thermodynamics, structure and dynamics. This strategy was applied to study the interaction between aminoglycoside N-(6')-acetyltransferase-Ii and one of its substrates, acetyl coenzyme A. It was found that homotropic allostery between the two active sites of the homodimeric enzyme is modulated by opposing mechanisms. One follows a classical Koshland-Némethy-Filmer (KNF) paradigm, whereas the other follows a recently proposed mechanism in which partial unfolding of the subunits is coupled to ligand binding. Competition between folding, binding and conformational changes represents a new way to govern energetic communication between binding sites.
- Published
- 2011
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