Germ warfare.

Background: The major bottleneck for determination of 3 D structures of proteins using X-rays is the production of diffraction quality crystals. Often proteins are subjected to chemical modification to improve the chances of crystallization Results: Here, we report the successful crystallization of a nuclease employing a reductive methylation protocol. The key to crystallization was the successful introduction of 44 new cohesive (NZ) CH...O contacts (3.2 – 3.7 Å) by the addition of 2 methyl groups to the side chain amine nitrogen (NZ) of 9 lysine residues of the nuclease. The new contacts dramatically altered the crystallization properties of the protein, resulting in crystals that diffracted to 1.2 Å resolution. Analytical ultracentrifugation analysis and thermodynamics results revealed a more compact protein structure with better solvent exclusion of buried Trp residues in the folded state of the methylated protein, assisting crystallization. Conclusion: In this study, introduction of novel cohesive (NZ)CH...O contacts by reductive methylation resulted in the crystallization of a protein that had previously resisted crystallization in spite of extensive purification and crystallization space screening. Introduction of (NZ)CH...O contacts could provide a solution to crystallization problems for a broad range of protein targets. Background The resolution and accuracy of the structural information provided by X-rays is unsurpassed when compared to other techniques employed to resolve the 3 D structure of proteins [1]. However, not all proteins can be made available for X-ray diffraction studies because of the inherent difficulty in obtaining single crystals of adequate size and quality [2]. A subset of such proteins that resist crystallization can be salvaged by employing a reductive methylation protocol [3-11]. A 26 kD nuclease from Pyrococcus furiosus used in the current study had resisted crystallization inspite of extensive purification and crystallization Published: 7 July 2007 BMC Structural Biology 2007, 7:46 doi:10.1186/1472-6807-7-46 Received: 10 February 2007 Accepted: 7 July 2007 This article is available from: http://www.biomedcentral.com/1472-6807/7/46 © 2007 Shaw et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.