Mechanistic Studies on Tryptophan Lyase (NosL): Identification of Cyanide as a Reaction Product

Abstract

Tryptophan lyase (NosL) catalyzes the formation of 3-methylindole-2-carboxylic acid and 3-methylindole from l-tryptophan. In this paper, we provide evidence supporting a formate radical intermediate and demonstrate that cyanide is a byproduct of the NosL-catalyzed reaction with l-tryptophan. These experiments require a major revision of the NosL mechanism and uncover an unanticipated connection between NosL and HydG, the radical SAM enzyme that forms cyanide and carbon monoxide from tyrosine during the biosynthesis of the metallo-cluster of the [Fe-Fe] hydrogenase.

Following the electrons: peculiarities in the catalytic cycles of radical SAM enzymes

“…New insight, however, comes from a report published while this Viewpoint was under review and suggests that cyanide is produced rather than formaldehyde and ammonia. 52 If correct, this would make the NosL-catalyzed reaction an oxidation of tryptophan such that the net reduction of SAM is in fact the expected outcome as discussed in the previous section.…”

“…The NosL reaction may actually be an oxidation yielding cyanide (HC(II)≡N) as a product rather than formaldehyde and ammonia. 52…”

New developments in RiPP discovery, enzymology and engineering

“…344 In recent years, NosL has been extensively studied, revealing that the reaction involves a remarkable intramolecular migration of the L-Trp carboxylate to the C2 position of the indole ring, the removal of the Ca atom and the amino group, and leaving the former Cb as a methyl group. [344][345][346][347][348][349][350][351][352][353] The mechanism by which the enzyme achieves this challenging reaction has been extensively rened as work on NosL has progressed. At rst, the prevalence of tryptophanyl and tyrosyl radicals in biology led to the proposal that following canonical homolytic cleavage of SAM, the resulting 5 0 -dAc abstracts a hydrogen atom from the indole nitrogen of L-Trp, rather than from a carbon center.…”

The Promiscuous Activity of the Radical SAM Enzyme NosL toward Two Unnatural Substrates

“…[ 6‐8 ] Production of MIA from L ‐Trp involves a highly unusual carbon skeleton rearrangement, in which the carboxylate of L ‐Trp migrates to the indole C2 and the Cα‐N unit is released as a cyanide. [ 9‐12 ]…”