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Protein Crystallization in Restricted Geometry: Advancing Old Ideas for Modern Times in Structural Proteomics

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Structural Proteomics

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 426))

 

In the structural genomics period traditional methods for protein crystallization have been eclipsed by automation using batch or vapor diffusion equilibration to find conditions conducive for protein crystal growth. Although many globular and soluble proteins predominantly from prokaryotes have been crystallized and their structures solved by high throughput approaches, the remaining difficult proteins require more systematic and reflective methods combining miniaturization and integration of modern and traditional crystallography techniques. One of these conventional methods is growing crystals in restricted geometry, which is a historically well-known concept and a practical technique under-used by today's crystallographers. This chapter presents practical guidelines to use capillaries for microbatch crystallization screening and counter-diffusion crystallization as valuable techniques to obtain protein crystals in confined volumes. The emphasis in the authors' application is to perform broad-based screening with a microgram amount of protein, optimize crystal growth in a supersaturation gradient, and undergo in situ x-ray data analysis for x-ray crystallography without invasive manipulation. Applications and concepts presented here bring to light future prerequisites for the next generation of automation for structural genomics.

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Acknowledgments

The techniques reported here are derived from the collaborative effort between the University of Alabama in Huntsville and The Scripps Research Institute. Proof of concept of counter-diffusion crystallization for In situ crystallography was developed with Juan Manual García-Ruiz and José A. Gavira. The authors thank NSF (Alabama Structural Biology Consortium NSF-EPSCoR ) and NIH (NIH Roadmap award GM073197)

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Authors and Affiliations

  1. Hauptman-Woodward Medical Research Institute, Buffalo, New York, USA

    Joseph D. Ng, Raymond C. Stevens & Peter Kuhn

Authors
  1. Joseph D. Ng
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  2. Raymond C. Stevens
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  3. Peter Kuhn
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Editors and Affiliations

  1. School of Molecular and Microbial Sciences, The University of Queensland, Brisbane, Queensland, Australia

    Bostjan Kobe

  2. School of Molecular and Microbial Biosciences, University of Sydney, Sydney, Australia

    Mitchell Guss

  3. School of Molecular and Microbial Sciences and Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, Australia

    Thomas Huber

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© 2008 Humana Press, a part of Springer Science+Business Media, LLC

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Ng, J.D., Stevens, R.C., Kuhn, P. (2008). Protein Crystallization in Restricted Geometry: Advancing Old Ideas for Modern Times in Structural Proteomics. In: Kobe, B., Guss, M., Huber, T. (eds) Structural Proteomics. Methods in Molecular Biology™, vol 426. Humana Press. https://doi.org/10.1007/978-1-60327-058-8_23

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  • DOI: https://doi.org/10.1007/978-1-60327-058-8_23

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-809-6

  • Online ISBN: 978-1-60327-058-8

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