title: Preparation and crystallization of dynamic NF-kappa B.Ikappa B complexes.
abstract: The formation of single, well-diffracting crystals is a requirement for any molecular structure determination by x-ray crystallography. Crystallization of biological macromolecules can represent a significant obstacle when the subject exhibits internal flexibility or indiscriminate self-association. In such cases, the removal of inherently flexible regions and the addition of stabilizing ligands can improve the probability of crystal formation and ordered growth. We have applied these principles in order to form crystals of the Rel homology region of transcription factor NF-kappaB in complex with its inhibitors IkappaBalpha and IkappaBbeta. None of these molecules crystallizes in the absence of a binding partner. Recombinant overexpression of truncated IkappaBalpha required selection of the correct start site. NF-kappaB.IkappaBalpha complex crystals formed under relatively stringent conditions. NF-kappaB. IkappaBbeta complex crystals were formed by analogy to NF-kappaB. IkappaBalpha, although some modifications in purification and complex formation were necessary due to differences between the inhibitors.
authors.affiliation: Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, California 92037-0359, USA.
authors.forename: T
authors.initials: T
authors.lastname: Huxford
authors.forename: S
authors.initials: S
authors.lastname: Malek
authors.forename: G
authors.initials: G
authors.lastname: Ghosh
mesh_terms: D000818:Animals; D001665:Binding Sites; D003001:Cloning, Molecular; D003460:Crystallization; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D019281:Dimerization; D004926:Escherichia coli; D020672:I-kappa B Proteins; D051379:Mice; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606205:Nfkbia protein, mouse; D011994:Recombinant Proteins; D000072000:NF-KappaB Inhibitor alpha