3noj

Publication Abstract from PubMed

HMG/CHA aldolase from Pseudomonas putida F1 catalyzes the last step of the bacterial protocatechuate 4,5- cleavage pathway. The enzyme's preferred substrates are 2-keto-4-hydroxy acids with a 4-carboxylate substitution. The enzyme also exhibits oxaloacetate decarboxylation and pyruvate alpha-proton exchange activity. Sodium oxalate is a competitive inhibitor of the aldolase reaction. The pH dependence of kcat/Km and kcat for the enzyme is consistent with a single deprotonation with pKa values of 8.0 +/- 0.1 and 7.0 +/- 0.1 for free enzyme and enzyme substrate complex, respectively. The 1.8 A x-ray structure shows a four-layered alpha-beta-beta-alpha sandwich structure with the active site at the interface of two adjacent subunits of a hexamer; this fold resembles the RNAse E inhibitor, RraA, but is novel for an aldolase. The catalytic site contains a magnesium ion ligated by Asp124 as well as three water molecules bound by Asp102 and Glu199. A pyruvate molecule binds the magnesium ion through both carboxylate and keto oxygen atoms, completing the octahedral geometry. The carbonyl oxygen also forms a hydrogen bonds with the guanadinium group of Arg123, which site-directed mutagenesis confirms is essential for catalysis. A mechanism for HMG/CHA aldolase is proposed on the basis of the structure, kinetics, and previously established features of other aldolase mechanisms.

Structural and kinetic characterization of 4-hydroxy-4-methyl-2-oxoglutarate (HMG)/4-carboxy-4-hydroxy-2-oxoadipate (CHA) aldolase: a protocatechuate degradation enzyme evolutionarily convergent with the HpaI and DmpG pyruvate aldolases.,Wang W, Mazurkewich S, Kimber MS, Seah SY J Biol Chem. 2010 Sep 15. PMID:20843800^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.