Structure and Mechanism of Pseudomonas Aeruginosa PhzD, an Isochorismatase From the Phenazine Biosynthetic Pathway

Published

May 1, 2003

Author(s)

J F. Parsons, K Calabrese, Edward Eisenstein, Jane E. Ladner

Abstract

PhzD from Pseudomonas aeruginosa, is an isochorismatase involved in the synthesis of bioactive metabolites known as phenazines. Phenzines are antimicrobial compounds that provide Pseudomonas with a competitive advantage in certain environments and may be partly responsible for the persistence of Pseudomonas infections In vivo, PhzD catalyzes the hydrolysis of the vinyl ether functional group of 2-amino-2-deoxyisochorismate yielding pyruvate and trans-2,3-dihydro-3-hydroxy-anthranilic acid, which is further processed by other proteins in the phenazine pathway. PhzD also displays activity toward 4-amino-4-deoxychorismate and chorismate. Here we report the 1.5 crystal structure of native PhzD, and the 1.6 structure of the D38A mutant of PhzD in complex with isochorismate. The structures reveal that isochorismate binds to the PhzD active site in a trans-diaxial conformation with the re face of its methylene pyruvyl carbon adjacent to the side chain carboxylate of D38 when the essentially identical structures are superimposed. The proximity of D38 to the vinyl ether group suggests a mechanism in which D38 acts as a general acid in the protonation of the substrate forming a carbocation/oxocarbonium ion that is then rapidly hydrated to form hemiketal intermediate. The hemiketal then probably decomposes spontaneously to products. The structure of PhzD is remarkably similar to other structures from a subfamily of a/ -hydrolase enzymes that includes pyrazinamidase and N-carbamoylsarcosine amidohydrolase. PhzD however, catalyzes unrelated chemistry and lacks a nucleophilic cysteine found in its close structural relatives. The vinyl ether hydrolysis catalyzed by PhzD represents yet another example of the catalytic diversity seen in the a/ hydrolase family, whose members are also known to hydrolyze amides, phosphates, phosphonates, epoxides, and C-X bonds.

Citation

Biochemistry

Volume

42

Issue

No. 19

Pub Type

Journals

Keywords

crystal structure, enzyme, isochorismatase, phenazine pathway, PhzD, pseudomonas

Citation

Parsons, J. , Calabrese, K. , Eisenstein, E. and Ladner, J. (2003), Structure and Mechanism of Pseudomonas Aeruginosa PhzD, an Isochorismatase From the Phenazine Biosynthetic Pathway, Biochemistry (Accessed July 27, 2024)