Investigation of the catalytic mechanism of pyruvate phosphate dikinase

Conference · Fri May 01 04:00:00 EDT 1987 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:6285515

Wang, H C; Mehl, A; Pocalyko, D; Dunaway-Mariano, D

Previous studies have shown that the kinetic mechanism of pyruvate phosphate dikinase (PPDK) from B.symbiosus is bi(ATP,P/sub i/) bi(AMP,PP/sub i/) uni(pyruvate) uni(PEP). Recent studies have shown the same mechanism for the P. shermanii enzyme. Dead end inhibitor and alternate substrate studies indicate that ATP and P/sub i/ binding is random but PP/sub i/ is released from the enzyme before AMP. MnS binding to the enzyme was monitored with 9 and 35 GHz esr. The free enzyme binds one MnS /subunit at a high affinity site (Ki approx. 20 M) and a minimum of one MnS /subunit at a low affinity sites. Only one MnS /subunit binds at a high affinity site on the E x AMPPNP x P/sub i/ complex. Two possible chemical mechanisms have been considered. The first involves the partial reaction E x ATP x P/sub i/ E x ADP x PP/sub i/ EP x AMP x PP/sub i/ and the other E x ATP x P/sub i/ EPP x AMP x P/sub i/ EP x AMP x PP/sub i/. They have failed to detect enzyme bound ADP at equilibrium and have failed in their attempt to isolate EPP. They also attempted to distinguish between the two mechanisms with exchange Rxs. PPDK will not catalyze ( UC)-AMP in equilibrium AMPPNP exchange in the presence of P/sub i/, will not catalyze (TSP)-P/sub i/ in equilibrium PP/sub i/ exchange in the presence of AMPCPP, and will not catalyze ( UC)-AMP in equilibrium ATP exchange in the presence of methylene phosphate. Rapid quench techniques will be used to detect ADP or EPP formation.

• Research Organization: Univ. of Maryland, College Park
• OSTI ID: 6285515
• Report Number(s): CONF-870644-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 46:6
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ADP
AFFINITY
AMP
ATP
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
CARBON 14 COMPOUNDS
CARBOXYLIC ACIDS
DAYS LIVING RADIOISOTOPES
ENZYMES
ISOTOPE APPLICATIONS
ISOTOPES
KETO ACIDS
KINETICS
LABELLED COMPOUNDS
LIGHT NUCLEI
MANGANESE COMPOUNDS
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
OXYGEN COMPOUNDS
PHOSPHATES
PHOSPHORUS 32
PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
PYRUVIC ACID
RADIOISOTOPES
REACTION KINETICS
TRACER TECHNIQUES
TRANSFERASES
TRANSITION ELEMENT COMPOUNDS