Biochemical and biophysical investigation of the HalM2 lanthipeptide synthetase using mass spectrometry

Abstract

The rapid emergence of antimicrobial resistance in clinical settings has called for renewed efforts to discover and develop new antimicrobial compounds. Lanthipeptides present a promising, genetically encoded molecular scaffold for the engineering of structurally complex, biologically active peptides. These peptide natural products are constructed by enzymes (lanthipeptide synthetases) with relaxed substrate specificity that iteratively modify the precursor lanthipeptide to generate structures with defined sets of thioether macrocycles. The mechanistic features that guide the maturation of lanthipeptides into their proper, fully modified forms are obscured by the complexity of the multistep maturation and the large size and dynamic structures of the synthetases and precursor peptides. Over the past several years, our lab has been developing a suite of mass spectrometry-based techniques that are ideally suited to untangling the complex reaction sequences and molecular interactions that define lanthipeptide biosynthesis. This review focuses on our development and application of these mass spectrometry-based techniques to investigate the biochemical, kinetic, and biophysical properties of the haloduracin β class II lanthipeptide synthetase, HalM2.