Characterization of the capping layer of selenium nanoparticles synthesized by environmental bacterial strains and new hypothesis on selenium nanoparticles transport in Bacillus mycoides SeITE01

Selenium nanoparticles (SeNPs) are 10 to 400nm spheres composed of zero-valent selenium. SeNPs can be synthesized either chemically or biologically by microorganisms, plant extracts or enzymes. Biogenic SeNPs display a capping layer of organic molecules, which confer unique characteristics to such SeNPs, e.g. a major stability over time and a more efficient antimicrobial activity. Composition and role of the capping layer are mostly unknown and currently under investigation. In this study, environmental strains Bacillus mycoides SeITE01, Stenotrophomonas maltophilia SeITE02, Achromobacter sp. R2A, Ensifer sp. R2D and Lysinibacillus sp. R1E are considered, which are able to biosynthesize SeNPs. In the first section, SeNPs from the five bacteria are analyzed: microplate colorimetric assays are performed in order to quantify total carbohydrates, protein and lipids contents of such SeNPs capping layers. Moreover, SeNPs are treated with different protocols to remove part of the organic layer. Effect of such treatments on capping composition and SeNPs stability are studied and compared for all the five strains. In the second section, SeNPs produced by B. mycoides SeITE01 are analyzed from a proteomic point of view: biogenic SeNPs capping layer proteins are identified. Chemical SeNPs exposed to a SeITE01 cell free extract are analyzed as well. Identified proteins are compared in order to establish which proteins bind specifically biogenic SeNPs and are more probably involved in SeNPs formation. Finally, a model for SeNPs transport through SeITE01 cell wall is formulated, based on proteomic evidence. Native proteins activity assay and microscopy analysis are performed, in order to confirm the new model. In conclusion, studying the organic capping layer of biogenic SeNPs from different strains is of paramount importance to understand the effect of such molecules on SeNPs characteristics and formulate hypotheses on biosynthesis mechanism. SeITE01, SeITE02, R2A, R2D and R1E biosynthesized SeNPs show a different ratio of carbohydrates, proteins and lipids components of the capping layer and differently respond to treatments. Particularly, Gram-negative strains (SeITE02, R2A, R2D) show similar composition and respond to treatments in a similar fashion, while Gram-positive strains (SeITE01, R1E) show more variability. For SeITE01, proteomic and microscopy analyses led to a new model formulation for SeNPs transport outside the cell. Together with previous studies, this new hypothesis can contribute to a more complete vision of SeNPs synthesis in this aerobic strain.