Abstract：
A robust and stable carbonic anhydrase (CA) system is indispensable for effectively sequestering carbon dioxide to mitigate climate change. While microbial surface display technology has been employed to construct an economically promising cell-displayed CO2-capturing biocatalyst, the displayed CA enzymes were prone to inactivation due to their low stability in harsh conditions. Herein, drawing inspiration from biomineralized diatom frustules, we artificially introduced biosilica shell materials to the CA macromolecules displayed on Escherichia coli surfaces. Specifically, we displayed a fusion of CA and the diatom-derived silica-forming Sil3K peptide (CA-Sil3K) on the E. coli surface using the membrane anchor protein Lpp-OmpA linker. The displayed CA-Sil3K (dCA-Sil3K) fusion protein underwent a biosilicification reaction under mild conditions, resulting in nanoscale self-encapsulation of the displayed enzyme in biosilica. The biosilicified dCA-Sil3K (BS-dCA-Sil3K) exhibited improved thermal, pH, and protease stability and retained 63 % of its initial activity after ten reuses. Additionally, the BS-dCA-Sil3K biocatalyst significantly accelerated the CaCO3 precipitation rate, reducing the time required for the onset of CaCO3 formation by 92 % compared to an uncatalyzed reaction. Sedimentation of BS-dCA-Sil3K on a membrane filter demonstrated a reliable CO2 hydration application with superior long-term stability under desiccation conditions. This study may open new avenues for the nanoscale-encapsulation of enzymes with biosilica, offering effective strategies to provide efficient, stable, and economic cell-displayed biocatalysts for practical applications.
摘要：
强大而稳定的碳酸酐酶（CA）系统对于有效地隔离二氧化碳以减轻气候变化是必不可少的。虽然微生物表面展示技术已被用于构建经济上有前途的细胞展示CO2捕获生物催化剂，所展示的CA酶由于其在苛刻条件下的低稳定性而易于失活。在这里，从生物矿化硅藻细胞壳中汲取灵感，我们人工将生物二氧化硅壳材料引入到大肠杆菌表面显示的CA大分子中。具体来说,我们使用膜锚定蛋白Lpp-OmpA接头在大肠杆菌表面上展示了CA和硅藻衍生的形成二氧化硅的Sil3K肽(CA-Sil3K)的融合体。显示的CA-Sil3K（dCA-Sil3K）融合蛋白在温和条件下进行生物硅化反应，导致显示的酶在生物二氧化硅中的纳米级自封装。生物硅化的dCA-Sil3K（BS-dCA-Sil3K）表现出改善的热，pH值,和蛋白酶稳定性，并在十次重复使用后保留了其初始活性的63％。此外,BS-dCA-Sil3K生物催化剂显著加快了CaCO3的沉淀速度，与未催化的反应相比,CaCO3形成开始所需的时间减少92%。BS-dCA-Sil3K在膜滤器上的沉降证明了可靠的CO2水合应用，在干燥条件下具有出色的长期稳定性。这项研究可能会为生物二氧化硅对酶的纳米级封装开辟新的途径，提供有效的策略来提供有效的，稳定,和经济的细胞显示的生物催化剂的实际应用。
