The shells of the Pacific oyster Crassostrea gigas contain calcite crystals with three types of microstructures: prismatic, chalky, and foliated layers. Many shell matrix proteins were annotated from the shells of C. gigas; however, it is unclear which SMPs play important roles in their shell mineralization. The matrix proteins have never been reported from the chalky layer. In this study, we identified a chalky layer-derived EGF-like domain-containing protein (CgELC) from the chalky layer of C. gigas shells. The gene sequence of the CgELC was encoded under CGI_ 10,017,544 of the C. gigas genome database. Only peptide fragments in the N-terminal region of CGI_ 10,017,544 were detected by LC-MS/MS analyses, suggesting that CGI_ 10,017,544 was digested at the predicted protease digestion dibasic site by post-translational modification to become a mature CgELC protein. We produced three types of CgELC recombinant proteins, namely, the full length CgELC, as well as the N-terminal and C-terminal parts of CgELC (CgELC-N or -C, respectively), for in vitro crystallization experiments. In the presence of these recombinant proteins, the aggregation of polycrystalline calcite was observed. Some fibrous organic components seemed to be incorporated into the calcite crystals in the presence of the r-CgELC protein. We also noted different features in the crystallization between CgELC-N and CgELC-C; some crystals were inhibited crystal plane formation and contained many columnar prisms inside the crystals (CgELC-N) and formed numerous holes on their surfaces (CgELC-C). These results suggest that CgELC is involved in crystal aggregation and incorporated into calcite crystals.

太平洋牡蛎Crassostrea gigas的壳含有方解石晶体，具有三种微结构：棱柱状、白垩状和叶状层。许多壳基质蛋白是从C. gigas的壳中注释出来的；然而，尚不清楚哪些 SMP 在其壳矿化中起重要作用。从未报道过来自白垩层的基质蛋白。在这项研究中，我们从C. gigas壳的白垩层中鉴定了一种白垩层衍生的含有 EGF 样结构域的蛋白质 (CgELC) 。CgELC 的基因序列在C. gigas的 CGI_10,017,544 下编码基因组数据库。通过LC-MS/MS分析仅检测到CGI_10,017,544的N-末端区域中的肽片段，表明CGI_10,017,544在预测的蛋白酶消化二碱基位点处通过翻译后修饰被消化成为成熟的CgELC蛋白。我们生产了三种类型的 CgELC 重组蛋白，即全长 CgELC，以及 CgELC 的 N 端和 C 端部分（分别为 CgELC-N 或 -C），用于体外结晶实验。在这些重组蛋白的存在下，观察到多晶方解石的聚集。在 r-CgELC 蛋白的存在下，一些纤维状有机成分似乎被掺入到方解石晶体中。我们还注意到 CgELC-N 和 CgELC-C 之间结晶的不同特征；一些晶体的晶面形成受到抑制，晶体内部含有许多柱状棱柱（CgELC-N），并在其表面形成许多孔洞（CgELC-C）。这些结果表明 CgELC 参与晶体聚集并结合到方解石晶体中。