Kinetic and isotope partitioning studies on the NAD/sup +/-malic enzyme from Ascaris suum. [/sup 14/C tracer techniques]
Initial velocity studies in both the direction of malate decarboxylation and pyruvate carboxylation indicate that the NAD/sup +/-malic enzyme from Ascaris suum possesses a sequential kinetic mechanism. The V/sub f//V/sub r/ was 30. The K/sub m/ values for malate and NAD/sup +/, substrates of the forward reaction, were 1.9 +- 0.3 mM and 55 +- 7 ..mu..m, respectively. The K/sub m/ values for the substrates in the reverse reaction varied because of the inability to saturate the pyruvate or CO/sub 2/ binding sites. In general, the K/sub m/ for NADH was essentially the same as for NAD/sup +/, but the values for pyruvate and CO/sub 2/ ranged from 5 to 10 mM and 30 to 60 mM, respectively. All product inhibition studies resulted in linear noncompetitive patterns except for the nucleotides, which were competitive. Tartronate, oxalacetate, and other inhibitors competitive with respect to malate were used as dead-end inhibitors versus NAD/sup +/ and were noncompetitive. These results differed markedly from studies on other malic enzymes and suggested that the ascarid malic enzyme did not have an ordered mechanism, but exhibited some randomness in binding of malate and NAD/sup +/. Confirmation of random substrate binding was obtained by the method of isotope partitioning. Kinetically viable binary complexes were found to occur with both (/sup 14/C)malate and (/sup 14/C)NAD/sup +/ and the metal enzyme complex. NAD/sup +/ and malate dissociate from this complex 2.3 to 2.6 and 4.6 to 11.5 times faster, respectively, than the V/(E/sub t/).
- Research Organization:
- North Texas State Univ., Denton
- OSTI ID:
- 6654839
- Journal Information:
- J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 253:6; ISSN JBCHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ANIMALS
ASCARIS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BINDING ENERGY
BIOCHEMICAL REACTION KINETICS
CARBON 14
CARBON COMPOUNDS
CARBON DIOXIDE
CARBON ISOTOPES
CARBON OXIDES
CARBOXYLATION
CARBOXYLIC ACIDS
CHALCOGENIDES
CHEMICAL REACTIONS
COENZYMES
DECARBOXYLATION
ENERGY
ENZYMES
EVEN-EVEN NUCLEI
HELMINTHS
HYDROXY ACIDS
INVERTEBRATES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MALIC ACID
NAD
NADH2
NEMATODES
NUCLEI
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
OXIDES
OXYGEN COMPOUNDS
PARASITES
RADIOISOTOPES
REACTION KINETICS
TRACER TECHNIQUES
YEARS LIVING RADIOISOTOPES