Characterization of C-S Lyase from C. diphtheriae: A Possible Target for New Antimicrobial Drugs

May 22, 2018

Title

Characterization of C-S Lyase from C. diphtheriae: A Possible Target for New Antimicrobial Drugs

Author

Alessandra Astegno, Alejandro Giorgetti, Alessandra Allegrini, Barbara Cellini, Paola Dominici

Year

2013

Journal

BioMed Research International

Abstract

The emergence of antibiotic resistance in microbial pathogens requires the identification of new antibacterial drugs. The biosynthesis of methionine is an attractive target because of its central importance in cellular metabolism. Moreover, most of the steps in methionine biosynthesis pathway are absent in mammals, lowering the probability of unwanted side effects. Herein, detailed biochemical characterization of one enzyme required for methionine biosynthesis, a pyridoxal-5′-phosphate (PLP-) dependent C-S lyase from Corynebacterium diphtheriae, a pathogenic bacterium that causes diphtheria, has been performed. We overexpressed the protein in E. coli and analyzed substrate specificity, pH dependence of steady state kinetic parameters, and ligand-induced spectral transitions of the protein. Structural comparison of the enzyme with cystalysin fromTreponema denticola indicates a similarity in overall folding. We used site-directed mutagenesis to highlight the importance of active site residues Tyr55, Tyr114, and Arg351, analyzing the effects of amino acid replacement on catalytic properties of enzyme. Better understanding of the active site of C. diphtheriae C-S lyase and the determinants of substrate and reaction specificity from this work will facilitate the design of novel inhibitors as antibacterial therapeutics.

Instrument

FP-8200

Keywords

Fluorescence, Chemical stability, Ligand binding, Biochemistry, Medicinal