Carmel, G., Leichus, B., Cheng, X., Patterson, S. D., Mirza, U., Chait, B. T., Kuret, J. (March 1994) Expression, purification, crystallization, and preliminary x-ray analysis of casein kinase-1 from Schizosaccharomyces pombe. J Biol Chem, 269 (10). pp. 7304-9. ISSN 0021-9258 (Print)
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.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | Amino Acid Sequence Casein Kinases Catalysis Chromatography, Ion Exchange Cloning, Molecular Crystallography, X-Ray Escherichia coli/genetics Kinetics Molecular Sequence Data Mutation Protein Conformation Protein Kinases/chemistry/ genetics/isolation & purification Research Support, U.S. Gov't, P.H.S. Schizosaccharomyces/ enzymology Sequence Deletion Spectrum Analysis, Mass |
| Subjects: | Investigative techniques and equipment > cloning Investigative techniques and equipment > assays > cloning bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes > kinase Investigative techniques and equipment > spectroscopy > mass spectrometry Investigative techniques and equipment > x ray crystallography organism description > yeast |
| CSHL Authors: | |
| Communities: | CSHL labs > Stillman lab |
| Depositing User: | Matt Covey |
| Date: | 11 March 1994 |
| Date Deposited: | 07 May 2015 19:47 |
| Last Modified: | 07 May 2015 19:47 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/31457 |
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