Eubacterial proteins are synthesized with a formyl group at the N-terminus which is hydrolytically removed from the nascent chain by the mononuclear iron enzyme peptide deformylase. Catalytic efficiency strongly depends on the identity of the bound metal. We have determined by X-ray crystallography the Fe2+, Ni2+ and Zn2+ forms of the Escherichia coli enzyme and a structure in complex with the reaction product Met-Ala-Ser. The structure of the complex, with the tripeptide bound at the active site, suggests detailed models for the mechanism of substrate recognition and catalysis. Differences of the protein structures due to the identity of the bound metal are extremely small and account only for the observation that Zn2+ binds more tightly than Fe2+ or Ni2+. The striking loss of catalytic activity of the Zn2+ form could be caused by its reluctance to change between tetrahedral and five-fold metal coordination believed to occur during catalysis. N-terminal formylation and subsequent deformylation

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Peptide deformylase is an essential metalloenzyme required for the removal of the formyl group at the N terminus of nascent polypeptide chains in eubacteria. The Escherichia coli enzyme uses Fe 2؉ and nearly retains its activity on substitution of the metal ion by Ni 2؉ . We have solved the structure of the Ni 2؉ enzyme at 1.9-Å resolution by x-ray crystallography. Each of the three monomers in the asymmetric unit contains one Ni 2؉ ion and, in close proximity, one molecule of polyethylene glycol. Polyethylene glycol is shown to be a competitive inhibitor with a K I value of 6 mM with respect to formylmethionine under conditions similar to those used for crystallization. We have also solved the structure of the inhibitor-free enzyme at 2.5-Å resolution. The two structures are identical within the estimated errors of the models. The hydrogen bond network stabilizing the active site involves nearly all conserved amino acid residues and well defined water molecules, one of which ligates to the tetrahedrally coordinated Ni 2؉ ion.

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