title: Structure of N-formimino-L-glutamate iminohydrolase from Pseudomonas aeruginosa.
abstract: N-Formimino-l-glutamate iminohydrolase (HutF), from Pseudomonas aeruginosa with a locus tag of Pa5106 ( gi|15600299 ), is a member of the amidohydrolase superfamily. This enzyme catalyzes the deamination of N-formimino-l-glutamate to N-formyl-l-glutamate and ammonia in the histidine degradation pathway. The crystal structure of Pa5106 was determined in the presence of the inhibitors N-formimino-l-aspartate and N-guanidino-l-glutaric acid at resolutions of 1.9 and 1.4 Å, respectively. The structure of an individual subunit is composed of two domains with the larger domain folding as a distorted (β/α)8-barrel. The (β/α)8-barrel domain is composed of eight β-strands flanked by 11 α-helices, whereas the smaller domain is made up of eight β-strands. The active site of Pa5106 contains a single zinc atom that is coordinated by His-56, His-58, His-232, and Asp-320. The nucleophilic solvent water molecule coordinates with the zinc atom at a distance of 2.0 Å and is hydrogen bonded to Asp-320 and His-269. The α-carboxylate groups of both inhibitors are hydrogen bonded to the imidazole moiety of His-206, the hydroxyl group of Tyr-121, and the side chain amide group of Gln-61. The side chain carboxylate groups of the two inhibitors are ion-paired with the guanidino groups of Arg-209 and Arg-82. Computational docking of high-energy tetrahedral intermediate forms of the substrate, N-formimino-l-glutamate, to the three-dimensional structure of Pa5106 suggests that this compound likely undergoes a re-faced nucleophilic attack at the formimino group by the metal-bound hydroxide. A catalytic mechanism of the reaction catalyzed by Pa5106 is proposed.
authors.affiliation: Department of Chemistry and ‡Department of Biochemistry & Biophysics, Texas A&M University , College Station, Texas 77843, United States.
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Fedorov
authors.forename: Ricardo
authors.initials: R
authors.lastname: Martí-Arbona
authors.forename: Venkatesh V
authors.initials: VV
authors.lastname: Nemmara
authors.forename: Daniel
authors.initials: D
authors.lastname: Hitchcock
authors.forename: Elena V
authors.initials: EV
authors.lastname: Fedorov
authors.forename: Steven C
authors.initials: SC
authors.lastname: Almo
authors.forename: Frank M
authors.initials: FM
authors.lastname: Raushel
mesh_terms: D000581:Amidohydrolases / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism; D005971:Glutamates / Q000737:chemistry / Q000378:metabolism*; D008024:Ligands; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D015032:Zinc / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D005971:Glutamates; D008024:Ligands; C053697:N-formylglutamate; D000581:Amidohydrolases; D015032:Zinc