title: A spectroscopic investigation of the interaction between nitrogen monoxide and copper sites of the fungal laccase from Rigidoporus lignosus.
abstract: The interaction of NO, with the copper centres of the laccase secreted by Rigidoporus lignosus was studied under both aerobic or anaerobic conditions. The reduction of the T1 site was always observed, as detected by the disappearance of the characteristic optical band at 604 nm (T3 presents probably the same behaviour because of the decreasing of the band at 330 nm) and the absence of its characteristic EPR signal, while T2 undergoes an initial partial and transitory reduction, its EPR signal intensity totally restoring after 24 h interaction. Different magnetic parameters of the T2 site have been detected, evidencing an increase of the hyperfine coupling constant. Furthermore, the number of superhyperfine lines on the fourth line of T2 copper was also found to increase from seven in the native to nine in the NO-treated laccase, this fact implying the coordination of a nitrogenous species to the T2 site. It was also shown that nitrite can be a source of NO, thus, paralleling the behaviour of NO-donor molecules or NO gas, but after longer interaction times. The nitrogenous species coordinated to T2 site is probably NO2-, which arises indirectly by NO oxidation. In order to understand the mechanistic pathway of this interaction, some experiments were also carried out in the presence of azide to study the interaction of NO with this laccase having its trinuclear cluster blocked by the presence of an exogenous ligand as N3-. After the addition of NO-donor molecules to the azide-treated laccase, a new EPR signal appeared at low temperatures, which is ascribable to the partially reduced T3 site, while the T1 and T2 sites were found to be totally reduced. The mechanistic pathway of the NO interaction seems to proceed through the reduction of T1 and T3 copper sites, followed by the coordination of nitrogenous species to T2.
authors.affiliation: Dipartimento di Scienze Chimiche, Universita di Catania, Viale A. Doria 6, 95125 Catania, Italy. rbobomo@unict.it
authors.forename: R P
authors.initials: RP
authors.lastname: Bonomo
authors.forename: B M G
authors.initials: BM
authors.lastname: Castronovo
authors.forename: A M
authors.initials: AM
authors.lastname: Santoro
mesh_terms: D001487:Basidiomycota / Q000201:enzymology*; D001665:Binding Sites; D003300:Copper / Q000737:chemistry*; D007527:Isoenzymes; D042845:Laccase / Q000737:chemistry* / Q000378:metabolism; D009569:Nitric Oxide / Q000737:chemistry*
chemical_list: D007527:Isoenzymes; D009569:Nitric Oxide; D003300:Copper; D042845:Laccase