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Structural proteomics of an archaeon

Nature Structural Biology volume 7, pages 903–909 (2000)Cite this article

Abstract

A set of 424 nonmembrane proteins from Methanobacterium thermoautotrophicum were cloned, expressed and purified for structural studies. Of these, ∼20% were found to be suitable candidates for X-ray crystallographic or NMR spectroscopic analysis without further optimization of conditions, providing an estimate of the number of the most accessible structural targets in the proteome. A retrospective analysis of the experimental behavior of these proteins suggested some simple relations between sequence and solubility, implying that data bases of protein properties will be useful in optimizing high throughput strategies. Of the first 10 structures determined, several provided clues to biochemical functions that were not detectable from sequence analysis, and in many cases these putative functions could be readily confirmed by biochemical methods. This demonstrates that structural proteomics is feasible and can play a central role in functional genomics.

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Figure 1: M.th. target ORFs.
Figure 2: Histogram of the number of M.th. proteins at the end of each step of the cloning, expression, purification and sample screening process.
Figure 3: A decision tree discriminating between soluble and insoluble proteins.
Figure 4: Backbone ribbon representations of the first 10 protein structures.

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Acknowledgements

We are grateful to J. Reeve for providing M.th. ΔH chromosomal DNA. We thank technicians, A. Engel, S. Beasely, B. Le and summer students F. Hsu, A. Tuite, G. Minoo, S. Fung J. Loo, and H. Javidni for help with protein expression and purification. We thank L. Daniels for a gift of coenzyme F420, and A. Ayed for performing the EMSA of MTH1615. We acknowledge funding support from the Canadian funding organizations, MRC/CIHR (C.H.A, A.M.E, V.B., K.G.), NSERC (C.D.M), PENCE (L.P.M., E.F.P.) and US DOE contracts (M.A.K., J.R.C.), NIH and the Keck Foundation (M.G.), PNNL Laboratory Director's Research and Development funds (M.A.K.) and a Sloan Foundation-DOE fellowship (Y.K.). Part of the NMR work was performed at EMSL (a national scientific users facility sponsored by DOE Biological and Environmental Research) located at PNNL and operated by Battelle. X-ray data were collected at the Advanced Photon Source supported by the U.S. DOE, Basic Energy Sciences, Office of Science, and BioCARS Sector 14 supported by the NIH.

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Author notes

  1. Akil Dharamsi

    Present address: Integrative Proteomics Inc., Toronto, Ontario, Canada

  2. Dinesh Christendat, Adelinda Yee, Alan R. Davidson and Emil F. Pai: These two authors contributed equally to this work.

Authors and Affiliations

  1. Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, 610 University Ave, Toronto, M5G 2M9, Ontario, Canada

    Dinesh Christendat, Adelinda Yee, Akil Dharamsi, Alexei Savchenko, Valerie Booth, Vivian Saridakis, Ning Wu, Emil F. Pai, Aled M. Edwards & Cheryl H. Arrowsmith

  2. Department of Molecular Biophysics and Biochemistry and Computer Science, PO Box 208114, Yale University, New Haven , 06520, Connecticut, USA

    Yuval Kluger & Mark Gerstein

  3. Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratories, EMSL 2569 K8-98, Richland, 99352, Washington, USA

    John R. Cort & Michael A. Kennedy

  4. Departments of Biochemistry and Molecular Biology, Chemistry and the Biotechnology Laboratory, 2146 Health Sciences Mall, University of British Columbia, Vancouver, V6T 1Z3, British Columbia, Canada

    Cameron D. Mackereth & Lawrence P. McIntosh

  5. Biotechnology Research Institute, National Research Council of Canada, 6100 Royalmount Ave., Montreal , H4P 2R2, Quebec, Canada

    Irena Ekiel

  6. Department of Biochemistry and Montreal Joint Centre for Structural Biology, McGill University, 3655 Promenade Sir William Osler, Montreal, H3G 1Y6, Quebec, Canada

    Guennadi Kozlov & Kalle Gehring

  7. Department of Biochemistry, University of Toronto, 1 Kings College Circle, Toronto, M5S 1A8, Ontario, Canada

    Karen L. Maxwell, Alan R. Davidson & Emil F. Pai

  8. Department of Molecular and Medical Genetics, University of Toronto, 1 Kings College Circle, Toronto, M5S 1A8, Ontario, Canada

    Alan R. Davidson & Emil F. Pai

  9. Banting and Best Department of Medical Research, C.H. Best Institute, University of Toronto, 112 College St., Toronto, M5G 1L6, Ontario, Canada

    Aled M. Edwards

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  1. Dinesh Christendat
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Correspondence to Aled M. Edwards or Cheryl H. Arrowsmith.

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Christendat, D., Yee, A., Dharamsi, A. et al. Structural proteomics of an archaeon. Nat Struct Mol Biol 7, 903–909 (2000). https://doi.org/10.1038/82823

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