Grand Challenge in Biogeochemistry
EMSL Project ID
12493
Abstract
Do multiheme cytochromes associated with the outer membrane of metal reducing bacteria (mtrC in Shewanella oneidensis and omcB in Geobacter sulfurreducens, for example) transfer electrons to Fe(III) and Mn(IV) oxides and if so, by what mechanisms?The answer to this question has potential to greatly advance understanding of bioreductive geochemical processes that regulate contaminant migration in subsurface and sediment/water systems.
Knowledge of the genome of Shewanella oneidensis and Geobacter sulfurreducens and well developed genetic systems will greatly facilitate the proposed experimental investigations. The cloning and expression system developed for S. oneidensis as part of GtL projects will be leveraged to allow relatively rapid production of sufficient quantities of outer membrane protein mtrC for research. The structure of mtrC from Shewanella and omcB from Geobacter, which is expected to be different, will be determined by NMR spectroscopy and X-ray diffraction, and the biophysical properties of their heme centers characterized by EPR spectroscopy and other methods. Structural and biophysical information on mtrC and omcB and their heme groups will be used to develop molecular models to compute their conformation before and after surface engagement, and also to compute electron transfer rates and mechanisms with solid phase and soluble electron acceptors in different electrostatic and hydration environments as may occur on and within the outer membrane.
In vitro experiments will be performed with mtrC to understand the fundamental mechanisms of its interfacial engagement with mineral surfaces. MtrC, and other companion proteins as necessary such as mtrB and mtrA, either as free cytochromes or embedded in suitable simulants of the cell envelope (e.g., micelles), will be used with artificial electron donors to probe the formation of electron transfer precursor surface complexes and rates of electron transfer to Fe(III) and Mn(IV) sites in oxide surfaces. Epitaxially-grown thin oxide films and mineral electrodes will be used to investigate the metalloprotein electrochemistry, surface coordination and site specificity, and the kinetics and thermodynamics of the cytochrome/mineral interfacial electron transfer reactions. The influence of mineral surface properties such as structure, defects, and the electronic properties thereof will be investigated and monitored with modern surface science methods. Integrated molecular modeling of the biochemical and mineral surface system components will provide insights on the roles of protein surface conformation and orientation, the reorganization energy, and electronic coupling between the cytochrome heme centers and the valence and conduction bands of the oxide.
In vivo experiments will exploit genome information to develop mutants of S. oneidensis and G. sulfurreducens where genes related to outer membrane cytochromes [e.g., their regulation, cellular location, secretion pathway, electron transfer partner(s)] are deleted or over-expressed for hypothesis evaluation. Mutants and the wild-types cultured to specific and reproducible metabolic states will be contacted with model and natural mineral surfaces to evaluate whole-cell electron transfer rates and controlling factors. New high resolution electron, soft X-ray, and force microscopy techniques will be developed to determine the architecture of the microbe-mineral interface and the molecular environment within the interface where the cytochrome-mineral interaction occurs.
Integration of these studies with the others described above will provide the first comprehensive understanding of how outer membrane cytochromes accomplish bio-directed electron transfer with mineral surfaces.
Project Details
Project type
Grand Challenge
Start Date
2005-02-03
End Date
2008-09-16
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
Fredrickson JK, and JM Zachara. 2008. "Electron Transfer at the Microbe-Mineral Interface: A Grand Challenge in Biogeochemistry." Geobiology 6(3):245-253.