title: S-Nitrosylation of human variant albumin Liprizzi (R410C) confers potent antibacterial and cytoprotective properties.
abstract: The S-nitrosylated forms of certain proteins such as albumin have been thought to be circulating endogenous reservoirs of nitric oxide (NO) and may have potential as NO donors in therapeutic applications. In this study, we investigated the characteristics of R410C, a genetic variant of human serum albumin with two free thiols at positions 34 (Cys-34) and 410 (Cys-410), as a NO carrier via S-nitroso formation. A biotin switch assay revealed that Cys-410 was more rapidly and efficiently nitrosylated than was Cys-34. Nitrosylation of Cys-410 introduced only small conformational changes in the protein, which were detected by far-UV circular dichroism but not by near-UV circular dichroism. In addition, both native R410C and S-nitrosylated R410C did not induce molecular heterogeneity through oligomerization. S-Nitrosylated R410C exhibited strong antibacterial activity against Salmonella typhimurium in vitro and suppressed apoptosis of U937 human promonocytic cells induced by Fas ligand. In a rat ischemia-reperfusion liver injury model, S-nitrosylated R410C treatment significantly reduced liver damage, as indicated by markedly decreased release of liver enzymes (aspartate aminotransferase and alanine aminotransferase). Pharmacokinetic analyses indicated retention of the S-nitroso moiety of S-nitrosylated R410C in circulation after i.v. injection, with an approximate half-life of 20.4 min in the mouse. These data suggest that R410C can be a useful NO carrier and can be regarded as a new class of S-nitrosylated proteins possessing antibacterial and cytoprotective properties with a circulation time sufficient for in vivo biological activity.
authors.affiliation: Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Ishima
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Sawa
authors.forename: Ulrich
authors.initials: U
authors.lastname: Kragh-Hansen
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Miyamoto
authors.forename: Sadaharu
authors.initials: S
authors.lastname: Matsushita
authors.forename: Takaaki
authors.initials: T
authors.lastname: Akaike
authors.forename: Masaki
authors.initials: M
authors.lastname: Otagiri
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D001665:Binding Sites; D002460:Cell Line; D018360:Crystallography, X-Ray; D019610:Cytoprotection; D004195:Disease Models, Animal; D006801:Humans; D007511:Ischemia / Q000188:drug therapy; D008099:Liver / Q000098:blood supply; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D008958:Models, Molecular; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics; D015427:Reperfusion Injury / Q000517:prevention & control; D026403:S-Nitrosothiols / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology / Q000627:therapeutic use; D012486:Salmonella typhimurium / Q000187:drug effects; D012709:Serum Albumin / Q000737:chemistry* / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D011994:Recombinant Proteins; D026403:S-Nitrosothiols; D012709:Serum Albumin