3NOJ

The structure of HMG/CHA aldolase from the protocatechuate degradation pathway of Pseudomonas putida

Summary for 3NOJ

• Entry DOI: 10.2210/pdb3noj/pdb
• Descriptor: 4-carboxy-4-hydroxy-2-oxoadipate aldolase/oxaloacetate decarboxylase, MAGNESIUM ION, PYRUVIC ACID, ... (5 entities in total)
• Functional Keywords: class ii aldolase, a-b-b-a sandwich, metalloprotein, lyase
• Biological source: Pseudomonas putida
• Total number of polymer chains: 1
• Total formula weight: 25976.89

Authors

Kimber, M.S.,Wang, W.,Mazurkewich, S.,Seah, S.Y.K. (deposition date: 2010-06-25, release date: 2010-09-15, Last modification date: 2023-11-15)

Primary citation

Wang, W.,Mazurkewich, S.,Kimber, M.S.,Seah, S.Y.
Structural and Kinetic Characterization of 4-Hydroxy-4-methyl-2-oxoglutarate/4-Carboxy-4-hydroxy-2-oxoadipate Aldolase, a Protocatechuate Degradation Enzyme Evolutionarily Convergent with the HpaI and DmpG Pyruvate Aldolases.
J.Biol.Chem., 285:36608-36615, 2010

PubMed Abstract: 4-Hydroxy-4-methyl-2-oxoglutarate/4-carboxy-4-hydroxy-2-oxoadipate (HMG/CHA) aldolase from Pseudomonas putida F1 catalyzes the last step of the bacterial protocatechuate 4,5-cleavage pathway. The preferred substrates of the enzyme are 2-keto-4-hydroxy acids with a 4-carboxylate substitution. The enzyme also exhibits oxaloacetate decarboxylation and pyruvate α-proton exchange activity. Sodium oxalate is a competitive inhibitor of the aldolase reaction. The pH dependence of k(cat)/K(m) and k(cat) for the enzyme is consistent with a single deprotonation with pK(a) values of 8.0 ± 0.1 and 7.0 ± 0.1 for free enzyme and enzyme substrate complex, respectively. The 1.8 Å x-ray structure shows a four-layered α-β-β-α 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 Asp-124 as well as three water molecules bound by Asp-102 and Glu-199'. A pyruvate molecule binds the magnesium ion through both carboxylate and keto oxygen atoms, completing the octahedral geometry. The carbonyl oxygen also forms hydrogen bonds with the guanadinium group of Arg-123, 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.

PubMed: 20843800
DOI: 10.1074/jbc.M110.159509

Experimental method

X-RAY DIFFRACTION (1.82 Å)