Catalytic properties of the archaeal S-adenosylmethionine decarboxylase from Methanococcus jannaschii.
S-Adenosylmethionine decarboxylase (AdoMetDC) is a pyruvoyl cofactor-dependent enzyme that participates in polyamine biosynthesis. AdoMetDC from the Archaea Methanococcus jannaschii is a prototype for a recently discovered class that is not homologous to the eucaryotic enzymes or to a distinct group of microbial enzymes. M. jannaschii AdoMetDC has a Km of 95 microm and the turnover number (kcat) of 0.0075 s(-1) at pH 7.5 and 22 degrees C. The turnover number increased approximately 38-fold at a more physiological temperature of 80 degrees C. AdoMetDC was inactivated by treatment with the imine reductant NaCNBH3 only in the presence of substrate. Mass spectrometry of the inactivated protein showed modification solely of the pyruvoyl-containing subunit, with a mass increase corresponding to reduction of a Schiff base adduct with decarboxylated AdoMet. The presteady state time course of the AdoMetDC reaction revealed a burst of product formation; thus, a step after CO2 formation is rate-limiting in turnover. Comparable D2O kinetic isotope effects of were seen on the first turnover (1.9) and on kcat/Km (1.6); there was not a significant D2O isotope effect on kcat, suggesting that product release is rate-limiting in turnover. The pH dependence of the steady state rate showed participation of acid and basic groups with pK values of 5.3 and 8.2 for kcat and 6.5 and 8.3 for kcat/Km, respectively. The competitive inhibitor methylglyoxal bis(guanylhydrazone) binds at a single site per (alphabeta) heterodimer. UV spectroscopic studies show that methylglyoxal bis(guanylhydrazone) binds as the dication with a 23 microm dissociation constant. Studies with substrate analogs show a high specificity for AdoMet.
Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111-2497, USA.
Zichun J
ZJ
Lu
George D
GD
Markham
D012437:Adenosylmethionine Decarboxylase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019843:Archaeal Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D002384:Catalysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D017017:Methanococcus / Q000201:enzymology*; D008935:Mitoguazone / Q000494:pharmacology
D019843:Archaeal Proteins; D004791:Enzyme Inhibitors; D012437:Adenosylmethionine Decarboxylase; D008935:Mitoguazone