The Involvement of Microbially Derived Extracellular Proteins in Nanoparticle Formation and Aggregation

While humans are newcomers to the field of nanoscience, microbes have been synthesizing functional nanoscale structures for billions of years. Bacteria have evolved the capability to produce proteins that can unite cellular processes with inorganic substrates, transfer electrons, template biomineralization, and facilitate adhesion. Biominerals are commonly nano-composite materials in which biomolecules such as proteins and/or polysaccharides act as a template to direct nanoparticle nucleation and growth. Understanding the capability of microbes to form nanoparticles and influence their reactive transport properties offers potential for bioremediation and materials synthesis applications. The identification of biomolecules and functional groups associated with biogenic nanoparticle formation in both environmental and laboratory systems is the objective of our research. Two such systems in which protein-nanoparticle interactions were studied are discussed. First, the biogenic reduction of selenium oxyanions to Se0 was studied in pure cultures of Veillonella atypica, Bacillus selenitireducens and Geobacter sulfurreducens. Biogenic Se0 nanostructures were observed as spherical, fibrillar, granular or amorphous aggregates, both in the cytoplasm or periplasmic space and extracellularly. These nanoparticles formed as protein-nanoparticle complexes that could be separated from the cells on the basis of density. A protein of ~39 kDa associated with biogenic nano-Se0 was recovered via polyacrylamide gel electrophoresis for characterization by MALDI-TOF mass spectrometry. Initial results suggest that this protein plays an integral, structural role in Se0 nanosphere formation. Second, the nanoparticulate products of bacterial sulfate reduction in a biofilm growing in minewater were investigated with multiple high- spatial resolution microanalyses. Biogenic zinc-sulfide nanoparticles exhibited evidence for rapid, highly efficient aggregation to form orders-of-magnitude larger spheroids. Analysis of these spheroids revealed a formative association between ZnS nanoparticles and microbially derived extracellular proteins. Direct protein extraction and isolation yielded a dominant ~37 kDa band for ongoing mass spectrometry characterization. Parallel experimental simulations of nanoparticle-amino acid interactions suggest an important role for cysteine-bearing proteins in promoting the aggregation of nanoparticulate metal-sulfides. Identification of microbial proteins that interact with natural or synthetic nanoparticles provides potential for synthesizing specific peptide sequences for technological and environmental applications, and sheds light into environmental nanoscale biomineralization processes that may significantly impact water quality.