Buey Rubén M, Monterroso Begoña, Menéndez Margarita, Diakun Greg, Chacón Pablo, Hermoso Juan Antonio, Díaz J Fernando
Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
J Mol Biol. 2007 Jan 12;365(2):411-24. doi: 10.1016/j.jmb.2006.09.091. Epub 2006 Oct 5.
Phosphocholine moieties decorating the pneumococcal surface are used as a docking station for a family of modular proteins, the so-called choline binding proteins or CBPs. Choline recognition is essential for CBPs function and may also be a determinant for their quaternary structure. There is little knowledge about modular arrangement or oligomeric structures in this family. Therefore, we have used the small angle X-ray scattering (SAXS) technique combined with analytical ultracentrifugation in order to model the three-dimensional envelope of two highly different CBPs: the phage encoded Cpl-1 lysozyme and the pneumococcal phosphorylcholine esterase Pce. Both enzymes have an N-terminal catalytic module and a C-terminal choline-binding module (CBM) that attaches them to the bacterial surface and comprises six and ten sequence repeats in Cpl-1 and Pce, respectively. SAXS experiments have shown an inherent conformational plasticity in Cpl-1 that accounts for the different relative position of these regions in the solution and crystal structures. Dimerization of Cpl-1 upon choline binding has been also visualised for the first time, and monomer-monomer interactions take place through the first CBR where a non-canonical choline binding site has now been identified. This mode of association seems to be independent of the absence or presence of the Cpl-1 catalytic module and reveals that the arrangement of the monomers differs from that previously found in the isolated CBM dimer of pneumococcal LytA amidase. In contrast, Pce displays the same modular disposition in the solution and crystal structures, and remains almost invariant upon choline binding. The present results suggest that protein dimerization and duplication of CBRs may be alternative but not equivalent ways of improving cell wall recognition by CBPs, since they provide different interaction geometries for choline residues present in (lipo)teichoic acids.
修饰肺炎球菌表面的磷酸胆碱部分用作一组模块化蛋白质（即所谓的胆碱结合蛋白或CBPs）的停靠站。胆碱识别对于CBPs的功能至关重要，也可能是其四级结构的决定因素。关于该家族中模块化排列或寡聚结构的知识很少。因此，我们使用小角X射线散射（SAXS）技术结合分析超速离心来模拟两种高度不同的CBPs的三维包络：噬菌体编码的Cpl-1溶菌酶和肺炎球菌磷酸胆碱酯酶Pce。这两种酶都有一个N端催化模块和一个C端胆碱结合模块（CBM），该模块将它们连接到细菌表面，在Cpl-1和Pce中分别包含六个和十个序列重复。SAXS实验表明Cpl-1具有内在的构象可塑性，这解释了这些区域在溶液和晶体结构中的不同相对位置。胆碱结合后Cpl-1的二聚化也首次被观察到，单体-单体相互作用通过第一个CBR发生，现在已经确定了一个非典型胆碱结合位点。这种结合模式似乎与Cpl-1催化模块的有无无关，并且揭示了单体的排列与先前在肺炎球菌LytA酰胺酶的分离CBM二聚体中发现的不同。相比之下，Pce在溶液和晶体结构中显示出相同的模块化排列，并且在胆碱结合后几乎保持不变。目前的结果表明，蛋白质二聚化和CBR的重复可能是CBPs改善细胞壁识别的替代但不等同的方式，因为它们为（脂）磷壁酸中存在的胆碱残基提供了不同的相互作用几何形状。
