title: Conformational heterogeneity within the Michaelis complex of lactate dehydrogenase.
abstract: A series of isotope edited IR measurements, both static as well as temperature jump relaxation spectroscopy, are performed on lactate dehydrogenase (LDH) to determine the ensemble of structures available to its Michaelis complex. There clearly has been a substantial reduction in the number of states available to the pyruvate substrate (as modeled by the substrate mimic, oxamate) and NADH when bound to protein compared to dissolved in solution, as determined by the bandwidths and positions of the critical C(2)═O band of the bound substrate mimic and the C(4)-H stretch of the NADH reduced nicotinamide group. Moreover, it is found that a strong ionic bond (characterized by a signature IR band discovered in this study) is formed between the carboxyl group of bound pyruvate with (presumably) Arg171, forming a strong "anchor" within the protein matrix. However, conformational heterogeneity within the Michaelis complex is found that has an impact on both catalytic efficiency and thermodynamics of the enzyme.
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York 10461, USA. call@aecom.yu.edu
authors.forename: Hua
authors.initials: H
authors.lastname: Deng
authors.forename: Dung V
authors.initials: DV
authors.lastname: Vu
authors.forename: Keith
authors.initials: K
authors.lastname: Clinch
authors.forename: Ruel
authors.initials: R
authors.lastname: Desamero
authors.forename: R Brian
authors.initials: RB
authors.lastname: Dyer
authors.forename: Robert
authors.initials: R
authors.lastname: Callender
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D007770:L-Lactate Dehydrogenase / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D009243:NAD / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D019289:Pyruvic Acid / Q000737:chemistry / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared; D014732:Vibration
chemical_list: D009243:NAD; D019289:Pyruvic Acid; D007770:L-Lactate Dehydrogenase