Conformational Dynamics of Biopolymers in the Course of Their Interaction: Multifaceted Approaches to the Analysis by the Stopped-Flow Technique with Fluorescence Detection

Abstract

This review deals with modern approaches to systematic research on molecular-kinetic mechanisms of damage recognition and removal by pro- and eukaryotic enzymes of DNA base excision repair. To this end, using DNA glycosylases from different structural families as an example—as well as apurinic/apyrimidinic endonuclease, which differs structurally and catalytically from DNA glycosylases—a comprehensive methodology is described in detail regarding studies on the mechanisms of action of DNA repair enzymes in humans and in Escherichia coli. This methodology is based on kinetic, thermodynamic, and mutational analyses of alterations in the conformation of molecules of an enzyme and of DNA during their interaction in real time. The described techniques can be used to analyze any protein–protein or protein–nucleic acid interactions.

Keywords

• enzymatic activity
• mechanism
• catalysis
• pre-steady-state kinetics
• thermodynamics
• mutational analysis