Jain Gaurav, Pendola Martin, Huang Yu-Chieh, Juan Colas Jose, Gebauer Denis, Johnson Steven, Evans John Spencer
Laboratory for Chemical Physics, Center for Skeletal and Craniofacial Biology, New York University , 345 East 24th Street, New York, New York 10010, United States.
Department of Chemistry, Physical Chemistry, Universität Konstanz , Universitätstrasse 10, Konstanz D-78457, Germany.
Biochemistry. 2017 Jul 18;56(28):3607-3618. doi: 10.1021/acs.biochem.7b00313. Epub 2017 Jul 7.
In the nacre or aragonitic layer of an oyster pearl, there exists a 12-member proteome that regulates both the early stages of nucleation and nanoscale-to-mesoscale assembly of nacre tablets and calcitic crystals from mineral nanoparticle precursors. Several approaches to understanding protein-associated mechanisms of pearl nacre formation have been developed, yet we still lack insight into how protein ensembles or proteomes manage nucleation and crystal growth. To provide additional insights, we have created a proportionally defined combinatorial model consisting of two pearl nacre-associated proteins, PFMG1 and PFMG2 (shell oyster pearl nacre, Pinctada fucata) whose individual in vitro mineralization functionalities are distinct from one another. Using scanning electron microscopy, atomic force microscopy, Ca(II) potentiometric titrations, and quartz crystal microbalance with dissipation monitoring quantitative analyses, we find that at 1:1 molar ratios, rPFMG2 and rPFMG1 co-aggregate in specific molecular ratios to form hybrid hydrogels that affect both the early and later stages of in vitro calcium carbonate nucleation. Within these hybrid hydrogels, rPFMG2 plays a role in defining protein co-aggregation and hydrogel dimension, whereas rPFMG1 defines participation in nonclassical nucleation processes; both proteins exhibit synergy with regard to surface and subsurface modifications to existing crystals. The interactions between both proteins are enhanced by Ca(II) ions and may involve Ca(II)-induced conformational events within the EF-hand rPFMG1 protein, as well as putative interactions between the EF-hand domain of rPFMG1 and the calponin-like domain of rPFMG2. Thus, the pearl-associated PFMG1 and PFMG2 proteins interact and exhibit mineralization functionalities in specific ways, which may be relevant for pearl formation.
在牡蛎珍珠的珍珠层或文石层中，存在一个由12种蛋白质组成的蛋白质组，它调节着成核的早期阶段以及从矿物纳米颗粒前体形成珍珠层片和方解石晶体的纳米尺度到中尺度的组装过程。已经开发了几种理解珍珠母形成过程中蛋白质相关机制的方法，但我们仍然缺乏对蛋白质集合体或蛋白质组如何控制成核和晶体生长的深入了解。为了提供更多见解，我们创建了一个按比例定义的组合模型，该模型由两种与珍珠母相关的蛋白质PFMG1和PFMG2（牡蛎珍珠母，合浦珠母贝）组成，它们各自的体外矿化功能彼此不同。通过扫描电子显微镜、原子力显微镜、Ca(II)电位滴定以及带有耗散监测定量分析的石英晶体微天平，我们发现，在1:1摩尔比下，重组PFMG2和重组PFMG1以特定的分子比例共同聚集形成混合水凝胶，这些水凝胶会影响体外碳酸钙成核的早期和后期阶段。在这些混合水凝胶中，重组PFMG2在定义蛋白质共聚集和水凝胶尺寸方面发挥作用，而重组PFMG1则决定参与非经典成核过程；两种蛋白质在对现有晶体的表面和亚表面修饰方面都表现出协同作用。两种蛋白质之间的相互作用因Ca(II)离子而增强，可能涉及Ca(II)诱导的EF手型重组PFMG1蛋白内的构象事件，以及重组PFMG1的EF手型结构域与重组PFMG2的钙调蛋白样结构域之间的假定相互作用。因此，与珍珠相关的PFMG1和PFMG2蛋白以特定方式相互作用并展现矿化功能，这可能与珍珠形成有关。
