The cysteinesulfenic Acid in NHase as catalytic nucleophile

Time-Resolved Crystallography of the Reaction Intermediate of Nitrile Hydratase: Revealing a Role for the Cysteinesulfenic Acid Ligand as a Catalytic Nucleophile.

Yamanaka, Y., Kato, Y., Hashimoto, K., Iida, K., Nagasawa, K., Nakayama, H., Dohmae, N., Noguchi, K., Noguchi, T., Yohda, M. and Odaka, M.

Angew. Chem. Int. Ed., (2015), 54: 10763–10767.

doi:10.1002/anie.201502731

The reaction mechanism of nitrile hydratase (NHase) was investigated using time-resolved crystallography of the mutant NHase, in which βArg56, strictly conserved and hydrogen bonded to the two post-translationally oxidized cysteine ligands, was replaced by lysine, and pivalonitrile was the substrate. The crystal structures of the reaction intermediates were determined at high resolution (1.2–1.3 Å). In combination with FTIR analyses of NHase following hydration in H218O, we propose that the metal-coordinated substrate is nucleophilically attacked by the O(SO−) atom of αCys114-SO−, followed by nucleophilic attack of the S(SO−) atom by a βArg56-activated water molecule to release the product amide and regenerate αCys114-SO−.

Unravelling the mechanism of NHase hydration

This paper (corresponding author, Masafumi Odaka) develops the knowledge around how a nitrile hydratase actually manages to stuff a water into a rather stable carbon-nitrogen triple bond. It also resulted in a rather nice suite of crystal structures- one parent structure (in pdb as 3A8O) and four single point mutants.

*Kinetic and structural studies on roles of the serine ligand and a strictly conserved tyrosine residue in nitrile hydratase - Yamanaka, Hashimoto, Ohtaki, Noguchi, Yohda and Odaka, in Journal of Biological Inorganic Chemistry (2010), doi: 10.1007/s00775-010-0632-3

This is an image produced using DeepView of 3A8O with the position of the iron labelled.