title: (NZ)CH...O contacts assist crystallization of a ParB-like nuclease.
abstract: 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 A) 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 A 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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China. neilshaw@moon.ibp.ac.cn <neilshaw@moon.ibp.ac.cn>
authors.forename: Neil
authors.initials: N
authors.lastname: Shaw
authors.forename: Chongyun
authors.initials: C
authors.lastname: Cheng
authors.forename: Wolfram
authors.initials: W
authors.lastname: Tempel
authors.forename: Jessie
authors.initials: J
authors.lastname: Chang
authors.forename: Joseph
authors.initials: J
authors.lastname: Ng
authors.forename: Xin-Yu
authors.initials: XY
authors.lastname: Wang
authors.forename: Sarah
authors.initials: S
authors.lastname: Perrett
authors.forename: John
authors.initials: J
authors.lastname: Rose
authors.forename: Zihe
authors.initials: Z
authors.lastname: Rao
authors.forename: Bi-Cheng
authors.initials: BC
authors.lastname: Wang
authors.forename: Zhi-Jie
authors.initials: ZJ
authors.lastname: Liu
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray / Q000379:methods; D019915:DNA Primase; D003851:Deoxyribonucleases / Q000737:chemistry*; D004706:Endodeoxyribonucleases / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry; D005090:Exodeoxyribonucleases / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D008745:Methylation; D008958:Models, Molecular; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020027:Pyrococcus furiosus / Q000201:enzymology*; D013816:Thermodynamics
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D019915:DNA Primase; D003851:Deoxyribonucleases; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C121791:dnaG protein, E coli; D008239:Lysine