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Examining Protein Crystallization Using Scanning Electron Microscopy

Published online by Cambridge University Press:  30 November 2012

Kathryn Gomery ,
Elaine C. Humphrey  and
Rodney Herring
Kathryn Gomery*
Affiliation:
University of Victoria, Department of Biochemistry and Microbiology, P.O. Box 3055 STN CSC, Victoria, BC V8W 3P6, Canada
Elaine C. Humphrey
Affiliation:
University of Victoria, Department of Mechanical Engineering, P.O. Box 3055 STN CSC, Victoria, BC V8W 3P6, Canada
Rodney Herring
Affiliation:
University of Victoria, Department of Mechanical Engineering, P.O. Box 3055 STN CSC, Victoria, BC V8W 3P6, Canada
*
*Corresponding author. E-mail: kgomery@uvic.ca
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Abstract

Elucidation of protein structure using X-ray crystallography relies on the quality of the crystal. Crystals suffer from many different types of disorder, some of which occur during crystal nucleation and early crystal growth. To date, there are few studies surrounding the quality and nucleation of protein crystals partly due to difficulties surrounding viewing biological samples at high resolution. Recent research has led our current understanding of nucleation to be a two-step mechanism involving the formation of nuclei from dense liquid clusters; it is still unclear whether nuclei first start as amorphous aggregate or as crystalline lattices. Our research examines this mechanism through the use of electron microscopy. Using scanning electron microscopy imaging of the protein crystal growth process, a stacking, spiraling manner of growth is observed. The tops of the pyramid-like tetragonal protein crystal structures measure ~0.2 μm across and contain ~125,000 lysozyme units. This noncrystalline area experiences strain due to growth of the protein crystal. Our work shows that it is possible to view detailed early stage protein crystal growth using a wet scanning electron microscopy technique, thereby overcoming the problem of viewing liquids in a vacuum.

Keywords

protein crystallographylysozymeSEMnucleation
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 19 , Issue 1 , February 2013 , pp. 145 - 149
Copyright
Copyright © Microscopy Society of America 2013

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References

Banfield, J.F., Welch, S.A., Zhang, H., Ebert, T.T. & Penn, R.L. (2000). Aggregation-based crystal growth and microstructure development in natural iron oxyhydroxide biomineralization products. Science 289(5480), 751–754.Google Scholar
Becker, R. (1940). On the formation of nuclei during precipitation. Proc Phys Soc 52, 71–76.CrossRefGoogle Scholar
Caylor, C.L., Dobrianov, I., Lemay, S.G., Kimmer, C., Kriminski, S., Finkelstein, K.D., Zipfel, W., Webb, W.W., Thomas, B.R., Chernov, A.A. & Thorne, R.E. (1999). Macromolecular impurities and disorder in protein crystals. Proteins 36(3), 270–281.Google Scholar
Chayen, N.E. (2005). Methods for separating nucleation and growth in protein crystallisation. Prog Biophys Mol Biol 88(3), 329–337.Google Scholar
Chernov, A.A. (2003). Protein crystals and their growth. J Struct Biol 142(1), 3–21.CrossRefGoogle ScholarPubMed
Chung, S.Y., Kim, Y.M., Kim, J.G. & Kim, Y.J. (2009). Multiphase transformation and Ostwald's rule of stages during crystallization of a metal phosphate. Nat Phys 5(1), 68–73.Google Scholar
Danilatos, G.D. (1994). Environmental scanning electron-microscopy and microanalysis. Mikrochim Acta 114, 143–155.Google Scholar
Donald, A.M. (2003). The use of environmental scanning electron microscopy for imaging wet and insulating materials. Nat Mater 2(8), 511–516.Google Scholar
Dunkle, J.A., Wang, L., Feldman, M.B., Pulk, A., Chen, V.B., Kapral, G.J., Noeske, J., Richardson, J.S., Blanchard, S.C. & Cate, J.H. (2011). Structures of the bacterial ribosome in classical and hybrid states of tRNA binding. Science 332(6032), 981–984.Google Scholar
Durbin, S.D. & Carlson, W.E. (1992). Lysozyme crystal-growth studied by atomic force microscopy. J Cryst Growth 122(1-4), 71–79.Google Scholar
Durbin, S.D. & Feher, G. (1990). Studies of crystal growth mechanisms of proteins by electron microscopy. J Mol Biol 212(4), 763–774.Google Scholar
Ebert, M., Inerle-Hof, M. & Weinbruch, S. (2002). Environmental scanning electron microscopy as a new technique to determine the hygroscopic behaviour of individual aerosol particles. Atmos Env 36(39-40), 5909–5916.Google Scholar
Feher, G. & Kam, Z. (1985). Nucleation and growth of protein crystals—General-principles and assays. Meth Enzymol 114, 77–112.Google Scholar
Frank, F.C. (1949). The influence of dislocations on crystal growth. Discuss Faraday Soc 5, 48–54.CrossRefGoogle Scholar
Gale, R. (1993). Growing protein crystals. In Crystallography Made Crystal Clear, pp. 37–45. San Diego, CA: Academic Press.Google Scholar
Galkin, O. & Vekilov, P.G. (2000). Control of protein crystal nucleation around the metastable liquid-liquid phase boundary. Proc Natl Acad Sci USA 97(12), 6277–6281.Google Scholar
Gourdon, P., Liu, X.Y., Skjorringe, T., Morth, J.P., Moller, L.B., Pedersen, B.P. & Nissen, P. (2011). Crystal structure of a copper-transporting PIB-type ATPase. Nature 475(7354), 59–64.Google Scholar
Helliwell, J.R. (2005). Protein crystal perfection and its application. Acta Crystallogr D 61(Pt 6), 793–798.Google Scholar
Kishita, K., Sakai, H., Tanaka, H., Saka, H., Kuroda, K., Sakamoto, M., Watabe, A. & Kamino, T. (2009). Development of an analytical environmental TEM system and its application. J Electron Microsc (Tokyo) 58(6), 331–339.Google Scholar
Leung, A.K., Nagai, K. & Li, J. (2011). Structure of the spliceosomal U4 snRNP core domain and its implication for snRNP biogenesis. Nature 473(7348), 536–539.Google Scholar
Mann, S., Moench, T.T. & Williams, R.J.P. (1984). A high-resolution electron-microscopic investigation of bacterial magnetite—Implications for crystal-growth. Proc R Soc Lond B Bio 221(1225), 385.Google Scholar
Manuel Garcia-Ruiz, J. (2003). Nucleation of protein crystals. J Struct Biol 142(1), 22–31.Google Scholar
Muscariello, L., Rosso, F., Marino, G., Giordano, A., Barbarisi, M., Cafiero, G. & Barbarisi, A. (2005). A critical overview of ESEM applications in the biological field. J Cell Physiol 205(3), 328–334.Google Scholar
Mutaftschiev, B. (1967). Formation de couches mixtes lors de la nucleation sur support etranger. J Phys Chem Solids S1, 599.Google Scholar
Park, Y.J., Lee, J.Y. & Kim, Y.T. (2006). In situ transmission electron microscopy study of the nucleation and grain growth of Ge2Sb2Te5 thin films. Appl Surf Sci 252(23), 8102–8106.CrossRefGoogle Scholar
Rossi, M.P., Ye, H.H., Gogotsi, Y., Babu, S., Ndungu, P. & Bradley, J.C. (2004). Environmental scanning electron microscopy study of water in carbon nanopipes. Nano Lett 4(5), 989–993.CrossRefGoogle Scholar
Saridakis, E. & Chayen, N.E. (2009). Towards a “universal” nucleant for protein crystallization. Trends Biotechnol 27(2), 99–106.Google Scholar
Turnbull, D. (1950). Formation of crystal nuclei in liquid metals. J Appl Phys 21(10), 1022–1028.Google Scholar
Vekilov, P.G. (2010a). Nucleation. Cryst Growth Des 10(12), 5007–5019.Google Scholar
Vekilov, P.G. (2010b). The two-step mechanism of nucleation of crystals in solution. Nanoscale 2(11), 2346–2357.Google Scholar
Volmer, M. & Adhikari, G. (1925). Experiments with crystal growth and decomposition. Z Physik 35(3), 170–176.CrossRefGoogle Scholar
Volmer, M. & Schultze, W. (1931). Condensation of crystals. Z Phys Chem A-Chem T 156(1), 1–22.Google Scholar
Wiencek, J.M. (1999). New strategies for protein crystal growth. Annu Rev Biomed Eng 1, 505–534.Google Scholar