title: Ibuprofen impairs allosterically peroxynitrite isomerization by ferric human serum heme-albumin.
abstract: Human serum albumin (HSA) participates in heme scavenging; in turn, heme endows HSA with myoglobin-like reactivity and spectroscopic properties. Here, the allosteric effect of ibuprofen on peroxynitrite isomerization to NO(3)(-) catalyzed by ferric human serum heme-albumin (HSA-heme-Fe(III)) is reported. Data were obtained at 22.0 degrees C. HSA-heme-Fe(III) catalyzes peroxynitrite isomerization in the absence and presence of CO(2); the values of the second order catalytic rate constant (k(on)) are 4.1 x 10(5) and 4.5 x 10(5) m(-1) s(-1), respectively. Moreover, HSA-heme-Fe(III) prevents peroxynitrite-mediated nitration of free added l-tyrosine. The pH dependence of k(on) (pK(a) = 6.9) suggests that peroxynitrous acid reacts preferentially with the heme-Fe(III) atom, in the absence and presence of CO(2). The HSA-heme-Fe(III)-catalyzed isomerization of peroxynitrite has been ascribed to the reactive pentacoordinated heme-Fe(III) atom. In the absence and presence of CO(2), ibuprofen impairs dose-dependently peroxynitrite isomerization by HSA-heme-Fe(III) and facilitates the nitration of free added l-tyrosine; the value of the dissociation equilibrium constant for ibuprofen binding to HSA-heme-Fe(III) (L) ranges between 7.7 x 10(-4) and 9.7 x 10(-4) m. Under conditions where [ibuprofen] is >>L, the kinetics of HSA-heme-Fe(III)-catalyzed isomerization of peroxynitrite is superimposable to that obtained in the absence of HSA-heme-Fe(III) or in the presence of non-catalytic HSA-heme-Fe(III)-cyanide complex and HSA. Ibuprofen binding impairs allosterically peroxynitrite isomerization by HSA-heme-Fe(III), inducing the hexacoordination of the heme-Fe(III) atom. These results represent the first evidence for peroxynitrite isomerization by HSA-heme-Fe(III), highlighting the allosteric modulation of HSA-heme-Fe(III) reactivity by heterotropic interaction(s), and outlining the role of drugs in modulating HSA functions. The present results could be relevant for the drug-dependent protective role of HSA-heme-Fe(III) in vivo.
authors.affiliation: Department of Biology and Interdepartmental Laboratory for Electron Microscopy, University Roma Tre, I-00146 Roma, Italy. ascenzi@uniroma3.it
authors.forename: Paolo
authors.initials: P
authors.lastname: Ascenzi
authors.forename: Alessandra
authors.initials: A
authors.lastname: di Masi
authors.forename: Massimo
authors.initials: M
authors.lastname: Coletta
authors.forename: Chiara
authors.initials: C
authors.lastname: Ciaccio
authors.forename: Gabriella
authors.initials: G
authors.lastname: Fanali
authors.forename: Francesco P
authors.initials: FP
authors.lastname: Nicoletti
authors.forename: Giulietta
authors.initials: G
authors.lastname: Smulevich
authors.forename: Mauro
authors.initials: M
authors.lastname: Fasano
mesh_terms: D000494:Allosteric Regulation; D006418:Heme / Q000737:chemistry*; D006801:Humans; D007052:Ibuprofen / Q000737:chemistry*; D007536:Isomerism; D007700:Kinetics; D008968:Molecular Conformation; D030421:Peroxynitrous Acid / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D012709:Serum Albumin / Q000737:chemistry*
chemical_list: D012709:Serum Albumin; D030421:Peroxynitrous Acid; D006418:Heme; D007052:Ibuprofen