Sites of binding and orientation in a four-location model for protein stereospecificity.
The stereospecificity of the enzyme isocitrate dehydrogenase was examined by steady-state kinetics and x-ray crystallography. The enzyme has the intriguing property that the apoenzyme in the absence of divalent metal showed a selectivity for the inactive l-enantiomer of the substrate isocitrate, whereas the enzyme containing magnesium showed selectivity for the physiologically active d-enantiomer. The hydrogen atom on the C2 carbon that is transferred during the reaction was, in both the d- and l-isocitrate complexes, in an orientation very close to that expected for delivery of a hydride ion to the cosubstrate NADP+. The beta-carboxylate that is eliminated as a CO2 molecule during the reaction occupied the same site on the protein in both the d- and l-isocitrate complexes. In addition, the C3 carbon was in the same protein site in both the d- and l-enantiomers. Only the fourth group, the OH atom, was in a very different position in the apo enzyme and in the metal-containing complexes. A four-location model is necessary to explain the enantiomeric specificity of IDH in contrast to the conventional three-point attachment model. The thermodynamic and kinetic ramifications of this model are explored.
Mesecar
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Center for Pharmaceutical Biotechnology and the Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, 60607, USA. mesecar@uic.edu
Koshland
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D001665:Binding Sites; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology; D007521:Isocitrate Dehydrogenase / Q000737:chemistry*; D007700:Kinetics; D008956:Models, Chemical; D008958:Models, Molecular; D011485:Protein Binding; D013237:Stereoisomerism; D013816:Thermodynamics
D007521:Isocitrate Dehydrogenase