Thesis
A chemical probe approach for studying protein redox dynamics in biofuel relevant cyanobacteria
Washington State University
Master of Science (MS), Washington State University
05/2016
Handle:
https://hdl.handle.net/2376/100337
Abstract
Biofuel research is inspired by the prospect of coercing microorganisms, with their skills in making high-energy molecules, into producing commercially relevant quantities of biofuel. Success in this venture would help ease our global dependency on non-renewable fossil fuels. Current approaches for tackling this technology include genetic annotation, pathway analysis and genetic modification; yet intracellular signaling mechanisms are often overlooked. Microbes utilize protein redox reactions for enacting fast and flexible metabolic change in response to their environment. Gaining insight into protein redox signaling would allow for a better understanding of the intricasies of microbial metabolism and assist in the development of strategies for exploiting microbial talents for biosynthesis. In order to study protein redox events, small molecule chemical probes were designed to cross cell membranes and covalently bind to reduced cysteine thiol moities. These probes were applied to two types of cyanobacteria in order to track protein redox reactions in cells as they were exposed to changes in nutrient availability. Probe labeled proteins were analyzed via SDS-PAGE, confocal microscopy, and high resolution LC-MS. This approach shed light on how reductant was partitioned within the cell, and resulted in the identification of redox sensitive proteins and the cysteine(s) responsible for this sensitivity. Cyanobacteria are a highly desirable microbial platform for biofuel production because they acquire energy through photosynthesis, sequester atmospheric CO2, and are metabolically diverse. Initial studies were focused on the cyanobacterium Synechococcus sp. PCC 7002. This organism can handle high levels of constant light, which for a phototroph means extended potential for production of organic molecules of interest that result from the conversion of light and atmospheric CO2 fixation. We limited and then starved Synechococcus sp. PCC 7002 cells of a carbon source to identify redox sensitive proteins affected by this nutrient perturbation. The second system we looked at was the cyanobacterium Cyanothece sp. ATCC 51142. Hydrogenases are sensitive to O2, yet Cyanothece can produce H2 concomitantly with photosynthesis, thereby puzzling scientists while inciting interest because of this organism’s unprecedented ability to reduce protons with photosynthetically derived electrons in the presence of O2.
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Details
- Title
- A chemical probe approach for studying protein redox dynamics in biofuel relevant cyanobacteria
- Creators
- Natalie Carol Sadler
- Contributors
- Allan S. Felsot (Chair)James R Pratt (Committee Member)Aaron Todd Wright (Committee Member) - Washington State University, Biological Systems Engineering, Department of
- Awarding Institution
- Washington State University
- Academic Unit
- Environment, School of the (CAHNRS)
- Theses and Dissertations
- Master of Science (MS), Washington State University
- Publisher
- Washington State University; [Pullman, Washington] :
- Number of pages
- 98
- Identifiers
- 99900525049401842
- Language
- English
- Resource Type
- Thesis