title: Application of the small-angle X-ray scattering technique for the study of two-step equilibrium enzyme-substrate interactions.
abstract: The small-angle x-ray scattering (SAXS) technique is used for the investigation of two-stage equilibrium macromolecular interactions of the enzyme-substrate type in solution. Experimental procedures and methods of analyzing the data obtained from SAXS have been elaborated. The algorithm for the data analysis allows one to determine the stoichiometric, equilibrium, and structural parameters of the enzyme-substrate complexes obtained. The thermodynamic characteristics for the formation of complexes of double-stranded oligonucleotide with Eco dam methyltransferase (MTase) have been determined and demonstrate a high cooperativity of MTase binding when the ternary complex containing the dimeric enzyme is formed. The structural parameters (Rg, Rc, semiaxes) have been determined for free enzyme and polynucleotides and of enzyme-substrate complexes, indicating structural rearrangements of the enzyme in the interaction with substrates.
authors.affiliation: Institute of Molecular Biology, Koltsovo, Novosibirsk Region, Russia.
authors.forename: F V
authors.initials: FV
authors.lastname: Tuzikov
authors.forename: V V
authors.initials: VV
authors.lastname: Zinoviev
authors.forename: V I
authors.initials: VI
authors.lastname: Vavilin
authors.forename: E G
authors.initials: EG
authors.lastname: Malygin
mesh_terms: D000465:Algorithms; D001483:Base Sequence; D001704:Biopolymers; D004798:Enzymes / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D008956:Models, Chemical; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D012542:Scattering, Radiation; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000378:metabolism; D013379:Substrate Specificity; D014965:X-Rays
chemical_list: D001704:Biopolymers; D004798:Enzymes; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; C058693:Dam methyltransferase; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); C499294:dam protein, E coli