title: Structure of nitrite bound to copper-containing nitrite reductase from Alcaligenes faecalis. Mechanistic implications.
abstract: The structures of oxidized, reduced, nitrite-soaked oxidized and nitrite-soaked reduced nitrite reductase from Alcaligenes faecalis have been determined at 1.8-2.0 A resolution using data collected at -160 degrees C. The active site at cryogenic temperature, as at room temperature, contains a tetrahedral type II copper site liganded by three histidines and a water molecule. The solvent site is empty when crystals are reduced with ascorbate. A fully occupied oxygen-coordinate nitrite occupies the solvent site in crystals soaked in nitrite. Ascorbate-reduced crystals soaked in a glycerol-methanol solution and nitrite at -40 degrees C remain colorless at -160 degrees C but turn amber-brown when warmed, suggesting that NO is released. Nitrite is found at one-half occupancy. Five new solvent sites in the oxidized nitrite bound form exhibit defined but different occupancies in the other three forms. These results support a previously proposed mechanism by which nitrite is bound primarily by a single oxygen atom that is protonable, and after reduction and cleavage of that N-O bond, NO is released leaving the oxygen atom bound to the Cu site as hydroxide or water.
authors.affiliation: Department of Biological Structure, School of Medicine, University of Washington, Seattle, Washington 98195-7420, USA.
authors.forename: M E
authors.initials: ME
authors.lastname: Murphy
authors.forename: S
authors.initials: S
authors.lastname: Turley
authors.forename: E T
authors.initials: ET
authors.lastname: Adman
mesh_terms: D000421:Alcaligenes / Q000201:enzymology*; D001665:Binding Sites; D004926:Escherichia coli; D008956:Models, Chemical; D008958:Models, Molecular; D009572:Nitrite Reductases / Q000737:chemistry / Q000378:metabolism*; D009573:Nitrites / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction
chemical_list: D009573:Nitrites; D009572:Nitrite Reductases; C052695:nitrite reductase, copper-containing