Structural and Functional Characterization of Malate Synthase G from Opportunistic Pathogen Pseudomonas aeruginosa

• Published in: Biochemistry (Easton) Vol. 56; no. 41; pp. 5539 - 5549
• Main Authors: McVey, Alyssa C, Medarametla, Prasanthi, Chee, Xavier, Bartlett, Sean, Poso, Antti, Spring, David R, Rahman, Taufiq, Welch, Martin
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 17.10.2017
• Subjects: Acetyl Coenzyme A - chemistry; Acetyl Coenzyme A - metabolism; Amino Acid Sequence; Apoenzymes - chemistry; Apoenzymes - genetics; Apoenzymes - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Binding Sites; Catalytic Domain; Computational Biology; Conserved Sequence; Crystallography, X-Ray; Expert Systems; Glyoxylates - chemistry; Glyoxylates - metabolism; Indoles - chemistry; Indoles - metabolism; Ligands; Magnesium - chemistry; Magnesium - metabolism; Malate Synthase - chemistry; Malate Synthase - genetics; Malate Synthase - metabolism; Models, Molecular; Molecular Docking Simulation; Molecular Structure; Protein Conformation; Protein Structure, Secondary; Pseudomonas aeruginosa - enzymology; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Sequence Alignment; Structural Homology, Protein
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/acs.biochem.7b00852

Pseudomonas aeruginosa is an opportunistic human pathogen recognized as a critical threat by the World Health Organization because of the dwindling number of effective therapies available to treat infections. Over the past decade, it has become apparent that the glyoxylate shunt plays a vital role in sustaining P. aeruginosa during infection scenarios. The glyoxylate shunt comprises two enzymes: isocitrate lyase and malate synthase isoform G. Inactivation of these enzymes has been reported to abolish the ability of P. aeruginosa to establish infection in a mammalian model system, yet we still lack the structural information to support drug design efforts. In this work, we describe the first X-ray crystal structure of P. aeruginosa malate synthase G in the apo form at 1.62 Å resolution. The enzyme is a monomer composed of four domains and is highly conserved with homologues found in other clinically relevant microorganisms. It is also dependent on Mg2+ for catalysis. Metal ion binding led to a change in the intrinsic fluorescence of the protein, allowing us to quantitate its affinity for Mg2+. We also identified putative drug binding sites in malate synthase G using computational analysis and, because of the high resolution of the experimental data, were further able to characterize its hydration properties. Our data reveal two promising binding pockets in malate synthase G that may be exploited for drug design.