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Purification, crystallization and preliminary X‐ray diffraction analysis of the kinase domain of human tousled‐like kinase 2
Author(s) -
Garrote Ana M.,
Redondo Pilar,
Montoya Guillermo,
Muñoz Inés G.
Publication year - 2014
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x14002581
Subject(s) - crystallization , protein kinase domain , diffraction , domain (mathematical analysis) , kinase , materials science , crystallography , chemistry , physics , optics , biochemistry , mathematics , gene , organic chemistry , mutant , mathematical analysis
Tousled‐like kinases (TLKs) are an evolutionarily conserved family of serine/threonine protein kinases involved in chromatin dynamics, including DNA replication and repair, transcription and chromosome segregation. The two members of the family reported in humans, namely TLK1 and TLK2, localize to the cell nucleus and are capable of forming homo‐ or hetero‐oligomers by themselves. To characterize the role of TLK2, its C‐terminal kinase domain was cloned and overexpressed in Escherichia coli followed by purification to homogeneity. Crystallization experiments in the presence of ATP‐γ‐S yielded crystals suitable for X‐ray diffraction analysis belonging to two different space groups: tetragonal I 4 1 22 and cubic P 2 1 3. The latter produced the best diffracting crystal (3.4 Å resolution using synchrotron radiation), with unit‐cell parameters a = b = c = 126.05 Å, α = β = γ = 90°. The asymmetric unit contained one protein molecule, with a Matthews coefficient of 4.59 Å 3 Da −1 and a solvent content of 73.23%.