Molecular properties of pyruvate bound to lactate dehydrogenase: A Raman spectroscopic study
Journal Article
·
· Proc. Natl. Acad. Sci. USA; (United States)
Lactate dehydrogenase catalyzes the addition of pyruvate to the four position of the nicotinamide ring of bound NAD+; this NAD-pyruvate adduct is bound tightly to the enzyme. We have used the adduct as a model for pyruvate in a competent ternary complex by comparing the Raman spectrum of the bound adduct with that for unliganded pyruvate. To understand the observed normal modes of pyruvate both as the bound adduct and in water, we have taken the Raman spectra of a series of /sup 13/C- and /sup 18/O-labeled pyruvates. We find that the carboxylate COO- moiety of pyruvate remains unprotonated at LDH's active site and forms an ion pair complex. The frequency of pyruvate's carbonyl C = O moiety shifts from 1710 cm-1 in water downward 34 cm-1 when pyruvate binds to LDH. This frequency shift corresponds to a ca. 34% polarization of the carbonyl bond, indicates a substantial interaction between the C = O group and enzyme, and is direct evidence for and is a measure of enzyme-induced electronic perturbation of the substrate needed for catalysis. This bond polarization is likely brought about by electrostatic interactions between the carbonyl moiety and the protonated imidazole group of His-195 and the guanidino group from Arg-109. We discuss how the data bear on the enzymatic chemistry of LDH.
- Research Organization:
- City Univ. of New York, NY (USA)
- OSTI ID:
- 5768301
- Journal Information:
- Proc. Natl. Acad. Sci. USA; (United States), Journal Name: Proc. Natl. Acad. Sci. USA; (United States) Vol. 86:12; ISSN PNASA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
BIOCHEMICAL REACTION KINETICS
CARBON 13
CARBON ISOTOPES
CARBOXYLIC ACIDS
CHEMICAL PROPERTIES
COENZYMES
ENZYMES
EVEN-EVEN NUCLEI
EVEN-ODD NUCLEI
HEMIACETAL DEHYDROGENASES
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
KETO ACIDS
KINETICS
LACTATE DEHYDROGENASE
LIGHT NUCLEI
NAD
NUCLEI
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
OXIDOREDUCTASES
OXYGEN 18
OXYGEN COMPOUNDS
OXYGEN ISOTOPES
PYRUVIC ACID
RAMAN SPECTRA
REACTION KINETICS
SPECTRA
STABLE ISOTOPES
TRACER TECHNIQUES
WATER
59 BASIC BIOLOGICAL SCIENCES
BIOCHEMICAL REACTION KINETICS
CARBON 13
CARBON ISOTOPES
CARBOXYLIC ACIDS
CHEMICAL PROPERTIES
COENZYMES
ENZYMES
EVEN-EVEN NUCLEI
EVEN-ODD NUCLEI
HEMIACETAL DEHYDROGENASES
HYDROGEN COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
KETO ACIDS
KINETICS
LACTATE DEHYDROGENASE
LIGHT NUCLEI
NAD
NUCLEI
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
OXIDOREDUCTASES
OXYGEN 18
OXYGEN COMPOUNDS
OXYGEN ISOTOPES
PYRUVIC ACID
RAMAN SPECTRA
REACTION KINETICS
SPECTRA
STABLE ISOTOPES
TRACER TECHNIQUES
WATER