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Lysozyme Crystallization

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Lysozyme Crystallization
The crystallization of lysozyme to model a step for protein structure determination
Asaad Ahmed
November 6, 2014
TA: Ujwal

Abstract: Discovery of new protein structures is essential for advancement in a variety of scientific fields. By determining structures of proteins, scientists are able to identify active sites, which can lead to accomplishments such as the development of pharmaceuticals that can enhance or inhibit certain proteins in a variety of signaling pathways. To determine protein structures, a general protocol of steps must be followed. Proteins must be purified, crystallized, and then through x-ray diffraction techniques a structure can be determined. However, the most crucial part of structure determination is the crystallization of a protein. A protein can be difficult to crystallize because an optimal environment must be found by adjusting temperature, pH, ion concentrations, and even concentration of the protein itself. To understand this process, lysozyme, a well-studied protein, was crystallized in a total of twelve differing environments. The amount of two different solutes, NaCl and polyethylene glycol (PEG), were adjusted in the environments and the resulting crystals were examined. It was found that increasing amounts of a solute and protein will encourage high amounts of smaller sized crystals.
Introduction:
The process to determine a protein’s structure starts with the purification of the desired protein. A protein must be placed in an environment with specific conditions that will enable it to be isolated from impurities; different techniques of chromatography are extremely useful in these situations. Purification techniques are utilized by crystallographers because pure proteins must be crystallized in order to determine structure. If proteins crystallize with impurities, it can ultimately affect crystal structure and give inaccurate x-ray diffraction results and hinder the process of identifying protein structures. Crystallographers



References: 1. Wlodawer A., Minor W., Dauter Z., Jaskolski M. (2013). Protein crystallography for aspiring crystallographers or how to avoid pitfalls and traps in macromolecular structure determination. FEBS J 280: 5705–5736. 2. Höltje JV. Lysozyme substrates. EXS. 1996;75:105–110. 3. Forsythe, E. L., Maxwell, D. L. & Pusey, M. (2002). Vapor diffusion, nucleation rates and the reservoir to crystallization volume ratio. Acta Cryst. D58, 1601–1605. 4. Alderton G., Fevold H.L. (1945). Direct crystallization of lysozyme from egg white and some crystalline salts of lysozyme. J. Biol. Chem., 157, 43. 5. Department of Biology Davidson College, Protein Crystallization. [Internet]. 2003. [2014 Nov 13]. Available from: http://www.bio.davidson.edu/molecular/MolStudents/spring2003/Kogoy/protein.html 6. University of Houston (UH) Department of Biology and Biochemistry. (2014). Biochemistry I Laboratory. Houston, Texas: University of Houston

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