Data for the co-expression and purification of human recombinant CaMKK2 in complex with calmodulin in Escherichia coli

Lisa Gerner; Steffi Munack; Koen Temmerman; Ann-Marie Lawrence-Dörner; Hüseyin Besir; Matthias Wilmanns; Jan Kristian Jensen; Bernd Thiede; Ian G Mills; Jens Preben Morth

doi:10.1016/j.dib.2016.06.033

Data for the co-expression and purification of human recombinant CaMKK2 in complex with calmodulin in Escherichia coli

Data Brief. 2016 Jun 29:8:733-40. doi: 10.1016/j.dib.2016.06.033. eCollection 2016 Sep.

Authors

Affiliations

¹ Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, Forskningsparken, University of Oslo and Oslo University Hospitals, 0349 Oslo, Norway.
² European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany; European Molecular Biology Laboratory Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
³ European Molecular Biology Laboratory Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg, Germany; Institute of Biochemistry, WWU Münster, Wilhelm-Klemm-Straße 2, 48149 Münster, Germany.
⁴ European Molecular Biology Laboratory Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
⁵ European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany.
⁶ Department of Molecular Biology and Genetics, Aarhus University, 8000 Aarhus, Denmark.
⁷ Department of Biosciences, University of Oslo, Norway; Biotechnology Centre of Oslo, University of Oslo, Norway.
⁸ Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, Forskningsparken, University of Oslo and Oslo University Hospitals, 0349 Oslo, Norway; Department of Molecular Oncology, Institute for Cancer Research, Oslo University Hospitals, Norway; Movember/Prostate Cancer UK Centre of Excellence for Prostate Cancer Research, Centre for Cancer Research and Cell Biology (CCRCB), Queen׳s University Belfast, UK.
⁹ Centre for Molecular Medicine Norway (NCMM), Nordic EMBL Partnership, Forskningsparken, University of Oslo and Oslo University Hospitals, 0349 Oslo, Norway; Institute for Experimental Medical Research, Oslo University Hospital, Norway.

Abstract

Calcium/calmodulin-dependent kinase kinase 2 (CaMKK2) has been implicated in a range of conditions and pathologies from prostate to hepatic cancer. Here, we describe the expression in Escherichia coli and the purification protocol for the following constructs: full-length CaMKK2 in complex with CaM, CaMKK2 'apo', CaMKK2 (165-501) in complex with CaM, and the CaMKK2 F267G mutant. The protocols described have been optimized for maximum yield and purity with minimal purification steps required and the proteins subsequently used to develop a fluorescence-based assay for drug binding to the kinase, "Using the fluorescent properties of STO-609 as a tool to assist structure-function analyses of recombinant CaMKK2" [1].

Keywords: AMPK, adenosine monophosphate-regulated kinase; CaM, calmodulin; CaMK, calcium/calmodulin-dependent kinase; CaMKK2; CaMKK2, calcium/calmodulin-dependent kinase kinase 2; CaMKK2:CaM, CaMKK2-CaM complex; Calmodulin; E. coli, Escherichia coli; Fermentation; IPTG, β-D-1-thiogalactopyranoside; LB, Luria broth; LDS-PAGE, lithium dodecyl sulphate–polyacrylamide gel electrophoresis; LIC, ligation-independent cloning; OD, optical density; PMSF, phenylmethylsulfonyl fluoride; SEC, size-exclusion chromatography; TEV, tobacco etch virus; aa, amino acid.