PNNL Team Determines Majority of “Conan the Bacterium’s” Proteome

The Science

Scientists at the Department of Energy’s Pacific Northwest National Laboratory have obtained the most complete analysis of the total set of proteins (the proteome) of any organism to date using the microbe Deinococcus radiodurans. This microbe has been called “Conan the Bacterium” by the media for its ability to withstand high doses of radiation and its astonishing DNA repair capabilities and has even been listed in the Guinness Book of World Records as the world’s toughest bacterium. This research could open up new opportunities to harness this remarkable bacterium for helping to clean up contaminated DOE sites. While the genomic DNA sequence of this microbe was published three years ago, that information by itself has not allowed researchers to understand the DNA damage repair system. Information was needed about which genes are expressed when the microbe is exposed to radiation, about the proteins that are expressed in increased amounts under the radiation stress, and about how these proteins are involved in the highly effective damage recognition and repair system of D. radiodurans. Researchers have developed a high-throughput methodology to characterize an organism’s dynamic proteome based on the combination of global enzymatic digestion, high-resolution liquid chromatographic separations, and analysis by Fourier transform ion cyclotron resonance mass spectrometry.

The Impact

Researchers developed a method for identifying peptides based on accurate mass tags (AMTs) for each protein expressed by a given organism. This approach provides greater sensitivity and dynamic range than previously achievable, more comprehensive coverage of expressed proteins, a basis for precise quantitation, and greater throughput for measurements of proteomes, because protein identification using AMTs circumvents the need for routine tandem MS. The initial study focused on the extremely radiation-resistant bacterium Deinococcus radiodurans, but the general approach can be used for any organism whose genome has been sequenced.

Summary

In a study published in the August 20 issue of the Proceedings of the National Academy of Sciences (PNAS), a team lead by Richard Smith identified more than 60% of the proteins in the possible set of proteins (the proteome) predicted for D. radiodurans from its genome, the most complete analysis of the proteome yet done by any group. The results include identification of proteins that are highly expressed by this microorganism under environmental stresses, and discovery of functional classifications for proteins that previously were uncharacterized. These unprecedented studies were conducted on the unique 11.4 tesla mass spectrometer at the Biological and Environmental Research’s Environmental Molecular Sciences Laboratory (EMSL).

Principal Investigator

Richard Smith
Environmental Molecular Sciences Laboratory

BER Program Manager

U.S. Department of Energy, Biological and Environmental Research (SC-33)
Biological Systems Science Division
[email protected]

Funding

Funding was provided by the Office of Science Natural and Accelerated Bioremediation Research and Genomes to Life programs. The PNAS research article is accompanied by a commentary written by Jan Mrázek of Stanford University.

The research was directed by Richard D. Smith of the EMSL, with his colleague Mary Lipton as the lead author on the article, and with collaborators from Louisiana State University and the Uniformed Services University of the Health Sciences.

References

Lipton, M. S., L. Paša-Tolić, G. A. Anderson, D. J. Anderson, D. L. Auberry, J. R. Battista, M. J. Daly, J. Fredrickson, K. K. Hixson, H. Kostandarithes, C. Masselon, L. M. Markillie, R. J. Moore, M. F. Romine, Y. Shen, E. Stritmatter, N. Tolić, H. R. Udseth, A. Venkateswaran, K.-K. Wong, R. Zhao, and R. D. Smith. 2002. “Global Analysis of the Deinococcus radiodurans Proteome by Using Accurate Mass Tags,” Proceedings of the National Academy of Sciences (PNAS) 99(17), 11049–54. DOI:10.1073/pnas.172170199.