An active-site lysine in avian liver phosphoenolpyruvate carboxykinase

Journal Article · Tue Sep 10 00:00:00 EDT 1991 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00100a018· OSTI ID:5820213

Guidinger, P F; Nowak, T ^[1]

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1. Univ. of Notre Dame, IN (United States)

The participation of lysine in the catalysis by avian liver phosphoenolpyruvate carboxykinase was studied by chemical modification and by a characterization of the modified enzyme. The rate of inactivation by 2,4-pentanedione is pseudo-first-order and linearly dependent on reagent concentration with a second-order rate constant of 0.36 {plus minus} 0.025 M{sup {minus}1} min{sup {minus}1}. Inactivation by pyridoxal 5{prime}-phosphate of the reversible reaction catalyzed by phosphoenolpyruvate carboxykinase follows bimolecular kinetics with a second-order rate constant of 7,700 {plus minus} 860 m{sup {minus}1} min{sup {minus}1}. Treatment of the enzyme or one lysine residue modified concomitant with 100% loss in activity. A stoichiometry of 1:1 is observed when either the reversible or the irreversible reactions catalyzed by the enzyme are monitored. A study of k{sub obs} vs pH suggests this active-site lysine has a pK{sub a} of 8.1 and a pH-independent rate constant of inactivation of 47,700 m{sup {minus}1} min{sup {minus}1}. Proton relaxation rate measurements suggest that pyridoxal 5{prime}-phosphate modification alters binding of the phosphate-containing substrates. {sup 31}P NMR relaxation rate measurements show altered binding of the substrates in the ternary enzyme {center dot}Mn{sup 2+}{center dot}substrate complex. Circular dichroism studies show little change in secondary structure of pyridoxal 5{prime}-phosphate modified phosphoenolpyruvate carboxykinase. These results indicate that avian liver phosphoenolpyruvate carboxykinase has one reactive lysine at the active site and it is involved in the binding and activation of the phosphate-containing substrates.

• OSTI ID: 5820213
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 30:36; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550600* -- Medicine
62 RADIOLOGY AND NUCLEAR MEDICINE
AMINO ACIDS
BARYONS
BIOCHEMICAL REACTION KINETICS
BODY
CARBOXYLIC ACIDS
DIGESTIVE SYSTEM
ELEMENTARY PARTICLES
ENZYMES
FERMIONS
GLANDS
HADRONS
INACTIVATION
ISOTOPES
KINETICS
LIGHT NUCLEI
LIVER
LYSINE
MAGNETIC RESONANCE
NUCLEAR MAGNETIC RESONANCE
NUCLEI
NUCLEONS
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANS
PHOSPHOENOLPYRUVATE
PHOSPHORUS 31
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
PROTEINS
PROTONS
REACTION KINETICS
RESONANCE
STABLE ISOTOPES
TRANSFERASES