title: Ligand binding to the FA3-FA4 cleft inhibits the esterase-like activity of human serum albumin.
abstract: The hydrolysis of 4-nitrophenyl esters of hexanoate (NphOHe) and decanoate (NphODe) by human serum albumin (HSA) at Tyr411, located at the FA3-FA4 site, has been investigated between pH 5.8 and 9.5, at 22.0°C. Values of Ks, k+2, and k+2/Ks obtained at [HSA] ≥ 5×[NphOXx] and [NphOXx] ≥ 5×[HSA] (Xx is NphOHe or NphODe) match very well each other; moreover, the deacylation step turns out to be the rate limiting step in catalysis (i.e., k+3 << k+2). The pH dependence of the kinetic parameters for the hydrolysis of NphOHe and NphODe can be described by the acidic pKa-shift of a single amino acid residue, which varies from 8.9 in the free HSA to 7.6 and 7.0 in the HSA:NphOHe and HSA:NphODe complex, respectively; the pK>a-shift appears to be correlated to the length of the fatty acid tail of the substrate. The inhibition of the HSA-Tyr411-catalyzed hydrolysis of NphOHe, NphODe, and 4-nitrophenyl myristate (NphOMy) by five inhibitors (i.e., diazepam, diflunisal, ibuprofen, 3-indoxyl-sulfate, and propofol) has been investigated at pH 7.5 and 22.0°C, resulting competitive. The affinity of diazepam, diflunisal, ibuprofen, 3-indoxyl-sulfate, and propofol for HSA reflects the selectivity of the FA3-FA4 cleft. Under conditions where Tyr411 is not acylated, the molar fraction of diazepam, diflunisal, ibuprofen, and 3-indoxyl-sulfate bound to HSA is higher than 0.9 whereas the molar fraction of propofol bound to HSA is ca. 0.5.
authors: Interdepartmental Laboratory of Electron Microscopy, Roma Tre University, Via della Vasca Navale 79, I-00146 Roma, Italy.
authors: Paolo
authors: P
authors: Ascenzi
authors: Department of Sciences, Roma Tre University, Viale Guglielmo Marconi 446, I-00146 Roma, Italy.
authors: Loris
authors: L
authors: Leboffe
authors: Department of Sciences, Roma Tre University, Viale Guglielmo Marconi 446, I-00146 Roma, Italy.
authors: Alessandra
authors: A
authors: di Masi
authors: Department of Sciences, Roma Tre University, Viale Guglielmo Marconi 446, I-00146 Roma, Italy.
authors: Viviana
authors: V
authors: Trezza
authors: Biomedical Research Division, Department of Theoretical and Applied Sciences, University of Insubria, Via Alberto da Giussano 12, I-21052 Busto Arsizio (VA), Italy.
authors: Gabriella
authors: G
authors: Fanali
authors: Department of Clinical Sciences and Translational Medicine, University of Roma "Tor Vergata", Via Montpellier 1, I-00133 Roma, Italy; Interuniversity Consortium for the Research on the Chemistry of Metals in Biological Systems, Via Celso Ulpiani 27, I-70126 Bari, Italy.
authors: Magda
authors: M
authors: Gioia
authors: Department of Clinical Sciences and Translational Medicine, University of Roma "Tor Vergata", Via Montpellier 1, I-00133 Roma, Italy; Interuniversity Consortium for the Research on the Chemistry of Metals in Biological Systems, Via Celso Ulpiani 27, I-70126 Bari, Italy.
authors: Massimo
authors: M
authors: Coletta
authors: Biomedical Research Division, Department of Theoretical and Applied Sciences, University of Insubria, Via Alberto da Giussano 12, I-21052 Busto Arsizio (VA), Italy.
authors: Mauro
authors: M
authors: Fasano
mesh_terms: D003975:Diazepam / Q000494:pharmacology; D004061:Diflunisal / Q000494:pharmacology; D004950:Esterases / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D004952:Esters / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis / Q000187:drug effects; D007052:Ibuprofen / Q000494:pharmacology; D008958:Models, Molecular; D015742:Propofol / Q000494:pharmacology; D012709:Serum Albumin / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D014443:Tyrosine / Q000037:antagonists & inhibitors* / Q000378:metabolism*
chemical_list: D004952:Esters; D012709:Serum Albumin; D014443:Tyrosine; D004061:Diflunisal; D004950:Esterases; D003975:Diazepam; D007052:Ibuprofen; D015742:Propofol