Analysis of enzyme-catalyzed nucleotide modification by aldose reductase

Conference · Fri May 01 00:00:00 EDT 1987 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:6079690

Grimshaw, C E

Homogeneous bovine kidney aldose reductase catalyzes two reactions in addition to the normal aldehyde-dependent oxidation of NADPH. First, adduct formation between the oxidized nucleotide and the oxidized substrate is observed during turnover due to initial formation of a reversible E:NADP/sup +/:R-CHO ternary complex, which subsequently reacts to give the covalent complex (E:NADP/sup +/-R-CHO). The reaction is enzyme-catalyzed with substantial enhancement of both the pseudo-first order rate constant and the overall K/sub eq/ relative to the reaction with free NADP/sup +/ in aqueous buffer. Analysis of the concentration dependence and time-course for reversible dead-end and covalent complex formation are described for several aldehyde and nucleotide substrates. Non-linear time courses for aldehyde reduction and substrate inhibition by the aldehyde substrate in initial velocity studies are completely accounted for by this mechanism, thereby eliminating a simple Dalziel-type explanation for the substrate activation by aldehyde which is also observed. Second, enzyme-catalyzed oxidation of NADPH occurs in the absence of aldehyde substrate with a rate equal to .03% of V/sub max/ for the normal reduction of glyceraldehyde. By 500 MHz /sup 1/H-NMR, the enzyme-catalyzed oxidation of (4-/sup 2/H)NADPH appears to be greater than 95% stereospecific. Spectroscopic evidence for a similar oxidation reaction is observed for the covalent E:NADP/sup +/-R-CHO adduct with glyceraldehyde, but not with glycolaldehyde.

• Research Organization: Research Institute of Scripps Clinic, La Jolla, CA
• OSTI ID: 6079690
• Report Number(s): CONF-870644-; TRN: 87-037118
• Journal Information: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 46:6; Conference: 78. annual meeting of the American Society of Biological Chemists conference, Philadelphia, PA, USA, 7 Jun 1987
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
NUCLEOTIDES
METABOLISM
OXIDOREDUCTASES
BIOCHEMICAL REACTION KINETICS
ALDEHYDES
CATTLE
COMPLEXES
DEUTERIUM
KIDNEYS
NADP
SUBSTRATES
TRACER TECHNIQUES
ANIMALS
BODY
COENZYMES
DOMESTIC ANIMALS
ENZYMES
HYDROGEN ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MAMMALS
NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
REACTION KINETICS
RUMINANTS
STABLE ISOTOPES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques