Vigil Toriana N, Rowson Mary-Jean C, Frost Abigail J, Janiga Abigail R, Berger Bryan W
Department of Chemical Engineering, University of Virginia, Charlottesville, Virginia 22903, United States.
Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia 22903, United States.
ACS Omega. 2025 Jan 6;10(1):334-343. doi: 10.1021/acsomega.4c06359. eCollection 2025 Jan 14.
Biomineralization is a green synthesis route for a variety of metal nanoparticles. Silicatein is a biomineralization protein originally found in marine sponge that converts inorganic precursors to metal oxide nanoparticles. In this work, we investigate the popular catalytic triad hypothesis and implement directed evolution with the aim to improve the solubility and kinetics of silicatein to enable increased nanoparticle synthesis. Site-directed mutagenesis with catalytic triad residues did not abolish biomineralization activity, aligning with the results seen in one previous study. Recombinant production of silicatein and mutants in following library generation and a survival screen yielded several mutant proteins with augmented biomineralization activity. Sequence analysis of these mutant proteins reveals multiple sequences within a single cell that contribute to enhanced biomineralization. Combined with the sequence analysis of silicateins from different marine sponges, these results suggest the protein is permissive to wide sequence variations and that multiple protein sequences act synergistically for enhanced biomineralization.
生物矿化是多种金属纳米颗粒的绿色合成途径。硅酸蛋白是最初在海洋海绵中发现的一种生物矿化蛋白，它能将无机前体转化为金属氧化物纳米颗粒。在这项工作中，我们研究了流行的催化三联体假说，并进行定向进化，目的是提高硅酸蛋白的溶解度和动力学，以增加纳米颗粒的合成。对催化三联体残基进行定点诱变并没有消除生物矿化活性，这与之前一项研究的结果一致。在生成文库并进行生存筛选后，重组生产硅酸蛋白及其突变体，得到了几种生物矿化活性增强的突变蛋白。对这些突变蛋白的序列分析揭示了单个细胞内有助于增强生物矿化的多个序列。结合来自不同海洋海绵的硅酸蛋白的序列分析，这些结果表明该蛋白允许广泛的序列变异，并且多个蛋白序列协同作用以增强生物矿化。
