Granier Thierry, Langlois d'Estaintot Béatrice, Gallois Bernard, Chevalier Jean-Marc, Précigoux Gilles, Santambrogio Paolo, Arosio Paolo
Unité de Biophysique Structurale, UMR CNRS 5471, Université Bordeaux I, Bât B8, avenue des Facultés, 33405 Talence Cedex, France.
J Biol Inorg Chem. 2003 Jan;8(1-2):105-11. doi: 10.1007/s00775-002-0389-4. Epub 2002 Sep 6.
The first ferritin structure refined at the atomic level has been achieved on recombinant mouse L-chain apoferritin (rMoLF) crystals. These latter diffract to 1.2 A resolution under cryogenic conditions. When cryo-cooling the sample, the thermal disorder usually observed at room temperature is reduced and the low-temperature structure reveals several details concerning the protein putative active sites and their properties. Within the pores built up by the molecular three-fold symmetry axes, the iron entry route to the ferritin cavity, residues H118, D131 and E134, exhibit alternate conformations associated with the binding of partially hydrated cadmium ions, a metal used as a crystallization agent. At the mineral ferrihydrite nucleation center, the electron density maps evidence the orientation of E57, E60, E61 and E64 glutamate side chains (whereas they were observed highly disordered in previous ferritin structures determined at room temperature) and allow a description of the site taking into account the binding geometry of four Cd(2+) ions. Moreover, the side chain of residue K140, lying in the vicinity of the ferrihydrite nucleation center, is shown to interact with residue E61. As previously highlighted, this observation confirms the importance of K140 in the rMoLF sequence, as being responsible for the low level of iron incorporation by mousel L-chain ferritin compared to human L-chain ferritin. Finally, the diffusion of small molecules within the ferritin cavity is illustrated here by the presence of ordered molecules of glycerol used as a cryo-protectant, which bind the inner cavity surface of the protein.
已在重组小鼠L链脱铁铁蛋白（rMoLF）晶体上实现了原子水平上精制的首个铁蛋白结构。后者在低温条件下可衍射至1.2埃分辨率。对样品进行低温冷却时，通常在室温下观察到的热无序现象会减少，低温结构揭示了有关蛋白质假定活性位点及其性质的若干细节。在由分子三重对称轴构成的孔内，铁进入铁蛋白腔的途径，即残基H118、D131和E134，呈现出与部分水合镉离子（用作结晶剂的一种金属）结合相关的交替构象。在矿物水铁矿成核中心，电子密度图证明了E57、E60、E61和E64谷氨酸侧链的取向（而在先前室温下测定的铁蛋白结构中观察到它们高度无序），并考虑到四个Cd(2+)离子的结合几何形状对该位点进行了描述。此外，位于水铁矿成核中心附近的残基K140的侧链显示与残基E61相互作用。如先前所强调的，这一观察结果证实了K140在rMoLF序列中的重要性，因为与人类L链铁蛋白相比，它导致小鼠L链铁蛋白的铁掺入水平较低。最后，用作冷冻保护剂的甘油有序分子的存在说明了小分子在铁蛋白腔内的扩散情况，这些甘油分子与蛋白质的内腔表面结合。
