Jain Gaurav, Pendola Martin, Huang Yu-Chieh, Gebauer Denis, 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.
Physical Chemistry, Department of Chemistry, Universität Konstanz , Universitätstrasse 10, D-78457 Konstanz, Germany.
Biochemistry. 2017 May 30;56(21):2663-2675. doi: 10.1021/acs.biochem.7b00083. Epub 2017 May 11.
In the purple sea urchin Strongylocentrotus purpuratus, the formation and mineralization of fracture-resistant skeletal elements such as the embryonic spicule require the combinatorial participation of numerous spicule matrix proteins such as SpSM50. However, because of its limited abundance and solubility issues, it has been difficult to pursue extensive in vitro biochemical studies of SpSM50 protein and deduce its role in spicule formation and mineralization. To circumvent these problems, we expressed a tag-free bacterial model recombinant spicule matrix protein, rSpSM50. Bioinformatics and biophysical experiments confirm that rSpSM50 is an intrinsically disordered, aggregation-prone C-type lectin-like domain-containing protein that forms dimensionally and internally heterogeneous protein hydrogels that control the in vitro mineralization process in three ways. The hydrogels (1) kinetically stabilize the aqueous calcium carbonate system against nucleation and thermodynamically destabilize the initially formed ACC in bulk solution, (2) promote and organize faceted single-crystal calcite and polycrystalline vaterite nanoparticles, and (3) promote surface texturing of calcite crystals and induce subsurface nanoporosities and channels within both calcite and vaterite crystals. Many of these features are also common to mollusk shell nacre proteins and the sea urchin spicule matrix glycoprotein, SpSM30B/C, and we conclude that rSpSM50 is a spiculogenesis hydrogelator protein that exhibits traits found in other calcium carbonate mineral-modification proteins.
在紫海胆（Strongylocentrotus purpuratus）中，诸如胚胎骨针等抗断裂骨骼元素的形成和矿化需要众多骨针基质蛋白（如SpSM50）的协同参与。然而，由于其丰度有限以及溶解性问题，对SpSM50蛋白进行广泛的体外生化研究并推断其在骨针形成和矿化中的作用一直很困难。为了规避这些问题，我们表达了一种无标签的细菌模型重组骨针基质蛋白rSpSM50。生物信息学和生物物理实验证实，rSpSM50是一种内在无序、易于聚集的含C型凝集素样结构域的蛋白质，它形成尺寸和内部均异的蛋白质水凝胶，以三种方式控制体外矿化过程。这些水凝胶（1）在动力学上稳定碳酸钙水溶液体系以防止成核，并在热力学上使本体溶液中最初形成的无定形碳酸钙不稳定，（2）促进并组织多面单晶方解石和多晶球霰石纳米颗粒，（3）促进方解石晶体的表面纹理化，并在方解石和球霰石晶体内部诱导亚表面纳米孔隙和通道。许多这些特征在软体动物贝壳珍珠层蛋白和海胆骨针基质糖蛋白SpSM30B/C中也很常见，我们得出结论，rSpSM50是一种骨针形成水凝胶蛋白，具有其他碳酸钙矿物改性蛋白中发现的特性。
