Tryptophan Lyase (NosL): Mechanistic Insights from Substrate Analogues and Mutagenesis

NosL is a member of a family of radical S-adenosylmethionine enzymes that catalyze the cleavage of the C-alpha-C-beta bond of aromatic amino acids. In this paper, we describe a set of experiments with substrate analogues and mutants for probing the early steps of the NosL mechanism. We provide biochemical evidence in support of the structural studies showing that the 5'-deoxyadenosyl radical abstracts a hydrogen atom from the amino group of tryptophan. We demonstrate that D-tryptophan is a substrate for NosL but shows relaxed regio control of the first beta-scission reaction. Mutagenesis studies confirm that Arg323 is important for controlling the regiochemistry of the beta-scission reaction and that Ser340 binds the substrate by hydrogen bonding to the indolic N1 atom.