Studies on Multi-nuclear Copper Enzymes and their Model Complexes

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The purpose of this project is to clarify the structure and function of Cu-containing nitrite reductase and nitrous oxide reductase.1) In order to shed light on the His255 and Asp98 residues around the type 2 Cu center in blue Achromobacter xylosoxidans nitrite reductase, two mutants were expressed in E. coil.The Asp98Ala and His255Ala mutants show the very low enzyme activities compared with the wild enzyme at pH 5.5, suggesting that these residues play a role of proton donation for the formation of water molecule.The Asp and His255 residues are also very important for the control of the intramolecular electron transfer rate from type 1 Cu to type 2.2) The nitrite reductase from Hyphomicrobium sp. A3151 has the two type 1 Cu's and one type 2 Cu, which is different from the other nitrite reductases containing type 1 Cu : type 2 Cu = 1 : 1. One of type 1 Cu(II) ions is blue and the other is green.3) The nitrous oxide reductase from Hyphomicrobium sp. A3151 is aerobically stable, and there are two forms, purple oxidized and blue reduced forms.4) The type 2 Cu(I) and (II) model complexes were prepared. The Cu(I) complex binding nitrite with nitrogen atom stoichiometrically gives NO gas in the presence of proton.