title: Expression, purification, crystallization, and preliminary x-ray analysis of casein kinase-1 from Schizosaccharomyces pombe.
abstract: The catalytic domain of Schizosaccharomyces pombe casein kinase-1 (the product of the cki1 gene) has been overexpressed in Escherichia coli, purified by chromatographic methods, characterized in vitro, and crystallized in the presence and absence of nucleotide substrate. The best crystals belong to the trigonal space group P3(1)21 or its enantiomorph, have unit cell parameters a = b = 79 A, c = 121 A, and diffract x-rays to 2.0-A resolution. Kinetic characterization of the purified catalytic domain and other C-terminal deletion mutants of Cki1 suggests that it is subject to two forms of regulation. One mechanism involves autophosphorylation, and results in a 4-fold decrease in the affinity for protein substrate. In contrast, truncation of intact Cki1 results in a 3-fold activation in its catalytic rate. This activation may arise from the removal of an inhibitory domain present in the intact enzyme.
authors.affiliation: Cold Spring Harbor Laboratory, New York 11724-2220.
authors.forename: G
authors.initials: G
authors.lastname: Carmel
authors.forename: B
authors.initials: B
authors.lastname: Leichus
authors.forename: X
authors.initials: X
authors.lastname: Cheng
authors.forename: S D
authors.initials: SD
authors.lastname: Patterson
authors.forename: U
authors.initials: U
authors.lastname: Mirza
authors.forename: B T
authors.initials: BT
authors.lastname: Chait
authors.forename: J
authors.initials: J
authors.lastname: Kuret
mesh_terms: D000595:Amino Acid Sequence; D047388:Casein Kinases; D002384:Catalysis; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D012568:Schizosaccharomyces / Q000201:enzymology*; D017384:Sequence Deletion
chemical_list: D011494:Protein Kinases; D047388:Casein Kinases