Berti Francesco, Gaggelli Elena, Guerrini Remo, Janicka Anna, Kozlowski Henryk, Legowska Anna, Miecznikowska Hanna, Migliorini Caterina, Pogni Rebecca, Remelli Maurizio, Rolka Krzysztof, Valensin Daniela, Valensin Gianni
Faculty of Chemistry, University of Wroclaw, F. Joliot-Curie 15, 50383 Wroclaw, Poland.
Chemistry. 2007;13(7):1991-2001. doi: 10.1002/chem.200601225.
Human prion protein (hPrP) fragments encompassing the 91-120 region, namely hPrP92-100 (SP1), hPrP106-113 (SP2), hPrP91-120 (LP1), and hPrP91-114 (LP2), were considered for delineation of the Cu(II)-binding site(s). NMR and EPR spectroscopy results obtained from LP1 or LP2 were compared with those obtained from SP1 and SP2. The coexistence of two binding sites, one centered at His96 and the other at His111, was evidenced and ratified by ESI mass spectrometry at low and high metal:peptide ratios. While room-temperature NMR spectroscopy data were consistent with the binding site centered on His111 being approximately fourfold stronger than that centered on His96, low-temperature EPR spectroscopy results yielded evidence for the opposite trend. This disagreement, which has also occurred in the literature, was clarified by temperature-dependent molecular dynamics runs that demonstrated Met112 approaching the metal at room temperature, a process that is expected to stabilize the His111-centered binding site through hydrophobic shielding of the metal coordination sphere.
为了确定铜（II）结合位点，研究了包含91-120区域的人朊病毒蛋白（hPrP）片段，即hPrP92-100（SP1）、hPrP106-113（SP2）、hPrP91-120（LP1）和hPrP91-114（LP2）。将从LP1或LP2获得的核磁共振（NMR）和电子顺磁共振（EPR）光谱结果与从SP1和SP2获得的结果进行比较。在低金属与肽比例和高金属与肽比例下，电喷雾电离质谱（ESI）证明并认可了两个结合位点的共存，一个以His96为中心，另一个以His111为中心。虽然室温核磁共振光谱数据表明，以His111为中心的结合位点比以His96为中心的结合位点强约四倍，但低温电子顺磁共振光谱结果显示出相反的趋势。文献中也出现过这种分歧，通过温度依赖性分子动力学模拟得以阐明，该模拟表明在室温下Met112靠近金属，这一过程有望通过对金属配位球的疏水屏蔽作用来稳定以His111为中心的结合位点。
