Catalytic mechanism of yeast adenosine 5'-monophosphate deaminase. Zinc content, substrate specificity, pH studies, and solvent isotope effects.
Adenosine 5'-monophosphate (AMP) deaminase from baker's yeast is an allosteric enzyme containing a single AMP binding site and two ATP regulatory sites per polypeptide [Merkler, D. J., & Schramm, V. L. (1990) J. Biol Chem. 265, 4420-4426]. The enzyme contains 0.98 +/- 0.17 zinc atom per subunit. The X-ray crystal structure for mouse adenosine deaminase shows zinc in contact with the attacking water nucleophile using purine riboside as a transition-state inhibitor [Wilson, D. K., Rudolph, F. B., & Quiocho, F. A. (1991) Science 252, 1278-1284]. Alignment of the amino acid sequence for yeast AMP deaminase with that for mouse adenosine deaminase demonstrates conservation of the amino acids known from the X-ray crystal structure to bind to the zinc and to a transition-state analogue. On the basis of these similarities, yeast AMP deaminase is also proposed to use a Zn(2+)-activated water molecule to attack C6 of AMP with the displacement of NH3. The pKm and pKi profiles for AMP and a competitive inhibitor overlap in a bell-shaped curve with pKa values of 7.0 and 7.4. This pattern is characteristic of a rapid equilibrium between AMP and the enzyme, thus confirming the rapid equilibrium random kinetic patterns [Merkler, D. J., Wali, A. S., Taylor, J., Schramm, V. L. (1989) J. Biol. Chem. 264, 21422-21430]. The Vmax of the reaction requires one unprotonated and one protonated group with pKa values of 6.4 +/- 0.2 and 7.7 +/- 0.3, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Merkler
D J
DJ
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York 10461.
Schramm
V L
VL
D000659:AMP Deaminase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D000249:Adenosine Monophosphate / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D003903:Deuterium; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D015394:Molecular Structure; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012997:Solvents; D013379:Substrate Specificity; D015032:Zinc / Q000032:analysis*
D012997:Solvents; D000249:Adenosine Monophosphate; D003903:Deuterium; D000659:AMP Deaminase; D015032:Zinc