title: A pivotal role for reductive methylation in the de novo crystallization of a ternary complex composed of Yersinia pestis virulence factors YopN, SycN and YscB.
abstract: Structural studies of a ternary complex composed of the Yersina pestis virulence factors YopN, SycN and YscB were initially hampered by poor solubility of the individual proteins. Co-expression of all three proteins in Escherichia coli yielded a well behaved complex, but this sample proved to be recalcitrant to crystallization. As crystallization efforts remained fruitless, even after the proteolysis-guided engineering of a truncated YopN polypeptide, reductive methylation of lysine residues was employed to alter the surface properties of the complex. The methylated complex yielded crystals that diffracted X-rays to a maximal resolution of 1.8 A. The potential utility of reductive methylation as a remedial strategy for high-throughput structural biology was further underscored by the successful modification of a selenomethionine-substituted sample.
authors.affiliation: Protein Engineering Section, Macromolecular Crystallography Laboratory, Center for Cancer Research, National Cancer Institute at Frederick, PO Box B, Frederick, MD 21702-1201, USA.
authors.forename: Florian D
authors.initials: FD
authors.lastname: Schubot
authors.forename: David S
authors.initials: DS
authors.lastname: Waugh
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008745:Methylation; D018832:Molecular Chaperones / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D017434:Protein Structure, Tertiary; D037521:Virulence Factors / Q000737:chemistry* / Q000378:metabolism*; D015010:Yersinia pestis / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D037521:Virulence Factors; C117911:YscB protein, Yersinia pestis; C070015:YopN protein, Yersinia