Fanali Gabriella, De Sanctis Giampiero, Gioia Magda, Coletta Massimo, Ascenzi Paolo, Fasano Mauro
Dipartimento di Biologia Strutturale e Funzionale, and Centro di Neuroscienze, Università dell'Insubria, Via Alberto da Giussano 12, 21052, Busto Arsizio (VA), Italy.
J Biol Inorg Chem. 2009 Feb;14(2):209-17. doi: 10.1007/s00775-008-0439-7. Epub 2008 Oct 21.
Human serum albumin (HSA) participates in heme scavenging, the bound heme turning out to be a reactivity center and a powerful spectroscopic probe. Here, the reversible unfolding of heme-HSA has been investigated by (1)H-NMR relaxometry, circular dichroism, and absorption spectroscopy. In the presence of 6 equiv of myristate (thus fully saturating all available fatty acid binding sites in serum heme-albumin), 1.0 M guanidinium chloride induces some unfolding of heme-HSA, leading to the formation of a folding intermediate; this species is characterized by increased relaxivity and enhanced dichroism signal in the Soret region, suggesting a more compact heme pocket conformation. Heme binds to the folding intermediate with K (d) = (1.2 +/- 0.1) x 10(-6) M. In the absence of myristate, the conformation of the folding intermediate state is destabilized and heme binding is weakened [K (d) = (3.4 +/- 0.1) x 10(-5) M]. Further addition of guanidinium chloride (up to 5 M) brings about the usual denaturation process. In conclusion, myristate protects HSA from unfolding, stabilizing a folding intermediate state in equilibrium with the native and the fully unfolded protein, envisaging a two-step unfolding pathway for heme-HSA in the presence of myristate.
人血清白蛋白（HSA）参与血红素清除，所结合的血红素成为反应中心和强大的光谱探针。在此，通过（1）H-NMR弛豫测量法、圆二色性和吸收光谱法研究了血红素-HSA的可逆去折叠。在存在6当量肉豆蔻酸盐的情况下（从而使血清血红素白蛋白中所有可用的脂肪酸结合位点完全饱和），1.0 M的氯化胍诱导血红素-HSA发生一些去折叠，导致形成一种折叠中间体；该物种的特征在于弛豫率增加以及在索雷特区域的二色性信号增强，这表明血红素口袋构象更紧凑。血红素以K（d）=（1.2±0.1）×10^（-6）M的解离常数与折叠中间体结合。在不存在肉豆蔻酸盐的情况下，折叠中间态的构象不稳定且血红素结合减弱[K（d）=（3.4±0.1）×10^（-5）M]。进一步添加氯化胍（高达5 M）会引发通常的变性过程。总之，肉豆蔻酸盐可保护HSA不发生去折叠，稳定一种与天然蛋白和完全去折叠蛋白处于平衡状态的折叠中间态，设想在肉豆蔻酸盐存在下血红素-HSA存在两步去折叠途径。
