Baroni S, Mattu M, Vannini A, Cipollone R, Aime S, Ascenzi P, Fasano M
Department of Chemistry IFM, University of Torino, Italy.
Eur J Biochem. 2001 Dec;268(23):6214-20. doi: 10.1046/j.0014-2956.2001.02569.x.
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.
血红素与人类血清白蛋白（HSA）结合赋予该蛋白质独特的光谱性质。本文报道了布洛芬和华法林对三价铁血红素 - 人类血清白蛋白（三价铁HSA - 血红素）以及亚硝基化亚铁血红素 - 人类血清白蛋白（亚铁HSA - 血红素 - NO）光谱性质的影响。三价铁HSA - 血红素是六配位的，血红素铁原子与His105和Tyr148结合。当药物与华法林的主要位点结合时，靠近血红素结合口袋埋藏的水分子被置换，导致发色团的电子吸收性质发生扰动，但不影响血红素铁的配位数或自旋状态，同时1H - NMR弛豫率降低。布洛芬和华法林与三价铁HSA - 血红素的华法林主要位点（对应于布洛芬的二级裂隙）结合的Kd值分别为5.4±1.1×10(-4) m和2.1±0.4×10(-5) m。布洛芬和华法林对三价铁HSA - 血红素的华法林主要裂隙的亲和力低于报道的药物与无血红素HSA结合的亲和力。因此，血红素与HSA结合的Kd值从无药物时的1.3±0.2×（10的-8次方）m增加到存在布洛芬和华法林时的1.5±0.2×（10的-7次方）m。亚铁HSA - 血红素 - NO是一种五配位的血红素铁体系。药物与亚铁HSA - 血红素 - NO的华法林主要位点结合会诱导向六配位血红素铁物种的转变，血红素铁原子与His105结合。值得注意的是，布洛芬的主要裂隙在功能和光谱上似乎与HSA的血红素位点解偶联。目前的结果为药物诱导的HSA变构平衡（a）的转变提供了明确的证据。
