Journal of Biological Chemistry
Volume 272, Issue 45, 7 November 1997, Pages 28455-28460
Journal home page for Journal of Biological Chemistry

PROTEIN CHEMISTRY AND STRUCTURE
Structure of Nitrite Bound to Copper-containing Nitrite Reductase from Alcaligenes faecalis: MECHANISTIC IMPLICATIONS*

https://doi.org/10.1074/jbc.272.45.28455Get rights and content
Under a Creative Commons license
open access

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 Å resolution using data collected at −160 °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 °C remain colorless at −160 °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.

Cited by (0)

*

This work was supported by National Institutes of Health Grant GM31770 (to E. T. A.) and by a Medical Research Council Post-doctoral Fellowship and a NSERC research grant (to M. E. P. M.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Current Address: Department of Biochemistry and Molecular Biology, University of British Columbia, 2146 Health Sciences Mall, Vancouver, V6T 1Z3 British Columbia, Canada.