Glassin, a protein occluded in biosilica of the hexactinellid sponge Euplectela, promotes silica formation from silicic acid at room temperature and neutral pH and is characterized by its primary structure which consists of a tandem repeat carrying three distinct domains, histidine and aspartic acid-rich (HD) domain, proline-rich (P) domain, and histidine and threonine-rich (HT) domain. The present study aims to clarify the domain responsible for the promotion of silica formation and to demonstrate usefulness of glassin and its domain as a tag for purification and immobilization of recombinant proteins. When each domain was mixed with silicic acid at neutral pH, silica was formed with HD domain as well as glassin, or a single repeat, but not with P or HT domain. Neither of amino or carboxy-terminal half of HD domain induced silica formation. The amount of silica formed with HD domain was significantly lower than that of glassin or a single repeat. HD domain fused with HT domain raised the amount of silica formed, while a HD domain fused with P domain, a mixture of HD and P domains, or a mixture of HD and HT domains has little effect on the promotion of silica formation. Collectively, a minimum sequence for promotion of silica formation is HD domain, whose activity can be enhanced by HT domain through a topological effect. In addition, practicality of glassin and HD domain was demonstrated by fusion of these sequences to green fluorescent protein which was successfully purified with Ni affinity chromatography and immobilized on silica.

格拉辛（一种蛋白被闭塞在海绵六倍体海绵Euplectela的生物二氧化硅中）在室温和中性pH下促进硅酸从二氧化硅形成，其主要结构由串联重复序列组成，该重复序列带有三个不同的域：组氨酸和富含天冬氨酸的（HD）域，富含脯氨酸的（P）域，和组氨酸和富含苏氨酸（HT）的域。本研究旨在阐明负责促进二氧化硅形成的结构域，并证明玻璃蛋白及其结构域作为纯化和固定重组蛋白的标签的有用性。当每个结构域在中性pH下与硅酸混合时，形成的二氧化硅具有HD结构域和玻璃蛋白，或单个重复序列，但不具有P或HT结构域。HD结构域的氨基或羧基末端一半均未诱导二氧化硅形成。具有HD域的形成的二氧化硅数量显着低于玻璃蛋白或单次重复的数量。与HT域融合的HD域增加了形成的二氧化硅的数量，而与P域，HD和P域的混合物或HD与HT域的混合物融合的HD域对促进二氧化硅形成的作用很小。总的来说，促进二氧化硅形成的最小顺序是HD结构域，其活性可以通过拓扑效应被HT结构域增强。另外，通过将这些序列与绿色荧光蛋白融合，证明了玻璃蛋白和HD结构域的实用性，绿色荧光蛋白通过Ni亲和色谱法成功纯化并固定在二氧化硅上。HD和P结构域的混合物，或HD和HT结构域的混合物对促进二氧化硅形成几乎没有影响。总的来说，促进二氧化硅形成的最小序列是HD结构域，其活性可以通过拓扑效应被HT结构域增强。另外，通过将这些序列与绿色荧光蛋白融合，证明了玻璃蛋白和HD结构域的实用性，绿色荧光蛋白通过Ni亲和色谱法成功纯化并固定在二氧化硅上。HD和P结构域的混合物，或HD和HT结构域的混合物对促进二氧化硅形成几乎没有影响。总的来说，促进二氧化硅形成的最小序列是HD结构域，其活性可以通过拓扑效应被HT结构域增强。另外，通过将这些序列与绿色荧光蛋白融合，证明了玻璃蛋白和HD结构域的实用性，绿色荧光蛋白通过Ni亲和色谱法成功纯化并固定在二氧化硅上。