Separation methods applicable to the evaluation of enzyme–inhibitor and enzyme–substrate interactions

Enzymes catalyze a rich variety of metabolic transformations, and do so with very high catalytic rates under mild conditions, and with high reaction regioselectivity and stereospecificity. These characteristics make biocatalysis highly attractive from the perspectives of biotechnology, analytical chemistry, and organic synthesis. This review, containing 128 references, focuses on the use of separation techniques in the elucidation of enzyme–inhibitor and enzyme–substrate interactions. While coverage of the literature is selective, a broad perspective is maintained. Topics considered include chromatographic methods with soluble or immobilized enzymes, capillary electrophoresis, biomolecular interaction analysis tandem mass spectrometry (BIA–MS), phage and ribosomal display, and immobilized enzyme reactors (IMERs). Examples were selected to demonstrate the relevance and application of these methods for determining enzyme kinetic parameters, ranking of enzyme inhibitors, and stereoselective synthesis and separation of chiral entities.