Elucidating the Mechanism behind the Bionanomanufacturing of Gold Nanoparticles Using Bacillus subtilis

Over the last two decades, gold nanoparticles (GNPs) have opened up numerous research and industrial opportunities in biomedical, optical, and electronic fields due to their size- and morphology-dependent properties [Grassian, V. H. Macromolecules 2008, 112(47), 18303-18313 and Nehl, C. L.; Hafner, J. H. J. Mater. Chem. 2008, 18(21), 2415-2419]. Therefore, green and efficient synthesis strategies providing precise control over size and morphology are desired. Since biological catalysts are known for the selectivity, efficiency, and environmentally friendly production of gold nanoparticles (referred to as bionanomanufacturing), they have been considered for GNP synthesis. However, the mechanism of how most of these biological entities produce GNPs has not been elucidated to date, limiting the industrial implementation of complex biological systems for nanoparticle synthesis. In this study, we investigated the mechanism of extracellular GNP production by Bacillus subtilis (B. subtilis). It is shown that B. subtilis releases vegetative catalase (Cat A) into the supernatant. Cat A from the supernatant and commercial catalase were employed to establish the mechanism of GNP formation. The bionanomanufactured GNPs were characterized using ultraviolet-visible (UV-vis) spectroscopy, transmission electron microscopy (TEM), and dynamic light scattering (DLS). Based on our results, we theorize that the mechanism of extracellular GNP production by B. subtilis Cat A involves (1) formation of gold-thiol bonds followed by (2) stabilization of GNPs with the denatured bacterial protein that serves as a capping agent. This research offers early insights into the gold-reducing mechanism occurring in the cell-free extract of B. subtilis, which can potentially lead to the design of protocols for the controlled production of GNPs with isolated enzymes at the industrial scale.