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Aspartate beta-decarboxylase from Alcaligenes faecalis: carbon-13 kinetic isotope effect and deuterium exchange experiments

Journal Article · · Biochemistry; (United States)
DOI:https://doi.org/10.1021/bi00328a004· OSTI ID:5220783
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The authors have measured the /sup 13/C kinetic isotope effect at pH 4.0, 5.0, 6.0, and 6.5 and in D/sub 2/O at pH 5.0 and the rate of D-H exchange of the alpha and beta protons of aspartic acid in D/sub 2/O at pH 5.0 for the reaction catalyzed by the enzyme aspartate beta-decarboxylase from Alcaligenes faecalis. The /sup 13/C kinetic isotope effect, with a value of 1.0099 +/- 0.0002 at pH 5.0, is less than the intrinsic isotope effect for the decarboxylation step, indicating that the decarboxylation step is not entirely rate limiting. The authors have been able to estimate probable values of the relative free energies of the transition states of the enzymatic reaction up to and including the decarboxylation step from the /sup 13/C kinetic isotope effect and the rate of D-H exchange of alpha-H. The pH dependence of the kinetic isotope effect reflects the pKa of the pyridine nitrogen of the coenzyme pyridoxal 5'-phosphate but not that of the imine nitrogen. A mechanism is proposed for the exchange of aspartate beta-H that is consistent with the stereochemistry suggested earlier.

Research Organization:
Univ. of Wisconsin, Madison
OSTI ID:
5220783
Journal Information:
Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 7; ISSN BICHA
Country of Publication:
United States
Language:
English

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Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
ASPARTIC ACID
BACTERIA
BARYONS
BIOCHEMICAL REACTION KINETICS
CARBON 13
CARBON ISOTOPES
CARBON-CARBON LYASES
CARBOXY-LYASES
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
COMPARATIVE EVALUATIONS
DECARBOXYLATION
DEUTERIUM
ELEMENTARY PARTICLES
ENZYME ACTIVITY
ENZYMES
EVEN-ODD NUCLEI
FERMIONS
HADRONS
HYDROGEN ISOTOPES
ION EXCHANGE
ISOTOPE EFFECTS
ISOTOPES
KINETICS
LIGHT NUCLEI
LYASES
MICROORGANISMS
NUCLEI
NUCLEONS
ODD-ODD NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PH VALUE
PROTONS
REACTION KINETICS
STABLE ISOTOPES
THERMODYNAMICS