title: Effect of ibuprofen and warfarin on the allosteric properties of haem-human serum albumin. A spectroscopic study.
abstract: Haem binding to human serum albumin (HSA) endows the protein with peculiar spectroscopic properties. Here, the effect of ibuprofen and warfarin on the spectroscopic properties of ferric haem-human serum albumin (ferric HSA-haem) and of ferrous nitrosylated haem-human serum albumin (ferrous HSA-haem-NO) is reported. Ferric HSA-haem is hexa-coordinated, the haem-iron atom being bonded to His105 and Tyr148. Upon drug binding to the warfarin primary site, the displacement of water molecules--buried in close proximity to the haem binding pocket--induces perturbation of the electronic absorbance properties of the chromophore without affecting the coordination number or the spin state of the haem-iron, and the quenching of the 1H-NMR relaxivity. Values of Kd for ibuprofen and warfarin binding to the warfarin primary site of ferric HSA-haem, corresponding to the ibuprofen secondary cleft, are 5.4 +/- 1.1 x 10(-4) m and 2.1 +/- 0.4 x 10(-5) m, respectively. The affinity of ibuprofen and warfarin for the warfarin primary cleft of ferric HSA-haem is lower than that reported for drug binding to haem-free HSA. Accordingly, the Kd value for haem binding to HSA increases from 1.3 +/- 0.2 x 10(-8) m in the absence of drugs to 1.5 +/- 0.2 x 10(-7) m in the presence of ibuprofen and warfarin. Ferrous HSA-haem-NO is a five-coordinated haem-iron system. Drug binding to the warfarin primary site of ferrous HSA-haem-NO induces the transition towards the six-coordinated haem-iron species, the haem-iron atom being bonded to His105. Remarkably, the ibuprofen primary cleft appears to be functionally and spectroscopically uncoupled from the haem site of HSA. Present results represent a clear-cut evidence for the drug-induced shift of allosteric equilibrium(a) of HSA.
authors.affiliation: Department of Chemistry IFM, University of Torino, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Baroni
authors.forename: M
authors.initials: M
authors.lastname: Mattu
authors.forename: A
authors.initials: A
authors.lastname: Vannini
authors.forename: R
authors.initials: R
authors.lastname: Cipollone
authors.forename: S
authors.initials: S
authors.lastname: Aime
authors.forename: P
authors.initials: P
authors.lastname: Ascenzi
authors.forename: M
authors.initials: M
authors.lastname: Fasano
mesh_terms: D000495:Allosteric Site; D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007052:Ibuprofen / Q000378:metabolism / Q000494:pharmacology*; D066298:In Vitro Techniques; D007700:Kinetics; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D012709:Serum Albumin / Q000737:chemistry* / Q000187:drug effects* / Q000378:metabolism; D013053:Spectrophotometry; D014859:Warfarin / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D046911:Macromolecular Substances; D012709:Serum Albumin; D006418:Heme; D014859:Warfarin; D007052:Ibuprofen