title: Distributive and directional behavior of lantibiotic synthetases revealed by high-resolution tandem mass spectrometry.
abstract: The lantibiotic synthetases LctM and HalM2 are bifunctional enzymes that catalyze both the dehydration of serine and threonine residues and the Michael-type additions of cysteine residues to the resulting dehydroamino acids in their substrate peptides. Using Fourier transform mass spectrometry to analyze these activities in vitro, the dehydration is shown to take place by a distributive mechanism, with build-up of intermediates observed in electrospray mass spectra. The cyclization activity of HalM2 was monitored through alkylation of free cysteines in intermediates, providing access to the regioselectivity of lanthionine ring formation using high-resolution tandem mass spectrometry. HalM2 is shown to catalyze the cyclization process in a largely N- to C-terminal directional fashion, forming a total of four lanthionine rings in its HalA2 substrate. These studies advance a model for lantibiotic production where substrate binding via an N-terminal leader results in dehydration and cyclization on similar time scales and with a high, though not strict, propensity for N-to-C directionality.
authors: Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, USA.
authors: M Violet
authors: MV
authors: Lee
authors: Leigh Anne Furgerson
authors: LA
authors: Ihnken
authors: Young Ok
authors: YO
authors: You
authors: Amanda L
authors: AL
authors: McClerren
authors: Wilfred A
authors: WA
authors: van der Donk
authors: Neil L
authors: NL
authors: Kelleher
mesh_terms: D000595:Amino Acid Sequence; D001430:Bacteriocins / Q000737:chemistry / Q000378:metabolism*; D055162:Biocatalysis; D003500:Cyclization; D007700:Kinetics; D008025:Ligases / Q000378:metabolism*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D053719:Tandem Mass Spectrometry
chemical_list: D001430:Bacteriocins; C082508:lacticin 481; D008025:Ligases