Identification of functional motifs for biomineralization in Phaeodactylum tricornutum silaffin

• Main Authors: Cheng, Yu-Heng, 鄭宇恆
• Other Authors: Lin, Han-Jia
• Format: Others
• Language: zh-TW
• Published: 2019
• Online Access: http://ndltd.ncl.edu.tw/handle/b59k2u

碩士 === 國立臺灣海洋大學 === 生命科學暨生物科技學系 === 107 === According to the patterns of frustule, diatoms can be roughly divided into two groups, centric diatoms and pinnate diatoms. Currently, the biomineralization process of frustule has been studied on Thalassiosira pseudonana, a centric diatom. Several proteins were identified as frustule synthesizing components, such as silaffins (silica affinity proteins). There are 4 silaffins were identified in T. pseudonana. The overall sequence identity of silaffins among different diatom species was low, but they still share the following features: containing a signal peptide, high composition of lysine and serine, and the pentalysine clusters. By using bioinformatics approach, we found a silaffin like gene (PtSil1) from a pinnate diatom, Phaeodactylum tricornutum. However, the repeated sequence of PtSil1 is different from the other silaffins in previous studies. Since this is the first silaffin identified from P. tricornutum, we aim to identify the functional motif for biosilicification of PtSil1. Based on the repeated sequence of PtSil1, we synthesized three peptides with different ratios of lysine, and tested the biosilicification activity respectively in vitro. Among them, the peptide PtS3 has higher activity than the previously reported peptide CfR5. In addition, the PtS3 also showed wider pH range for biosilicification. By using scanning electron microscopy (SEM), we found that PtS3 could enhance the formation of spherical silica nanoparticles. Moreover, the PtS3-fused green fluorescent protein (EGFP-S3) is able to facilitate the formation of spherical silica nanoparticle with green fluorescence. We also found that EGFP-S3 can resist from the denaturing effect of SDS after silica nanoparticle formation. In the future, we will apply this technology to protect commercial enzymes by fusing with the biosilicification peptide (PtS3).