Purification and crystallization of the N-terminal domain from the human doublecortin-like kinase.
The unique doublecortin-like tandem of two homologous domains is found in certain microtubule-associated proteins such as doublecortin (DCX) and doublecortin-like kinase (DCLK). It is responsible for interactions with tubulin/microtubules and regulates microtubule dynamics. Here, the expression and purification of the tandem from human DCLK (residues 49-280) and of the isolated domains (residues 49-154 and 176-280) and the successful crystallization of the N-terminal domain (N-DCLK) are reported. High-quality wild-type crystals were obtained and a complete native data set was collected to 1.5 A resolution. The crystals belong to space group C2, with unit-cell parameters a = 85.98, b = 29.62, c = 40.33 A, beta = 101.3 degrees. Crystals of SeMet-substituted N-DCLK (Leu120Met) were also obtained, but they exhibit the symmetry of space group P2(1), with unit-cell parameters a = 38.81, b = 29.43, c = 40.1 A, beta = 115.7 degrees.
Kim
Myung Hee
MH
Department of Molecular Physiology and Biological Physics and the Cancer Center, University of Virginia, PO Box 800736, Charlottesville, Virginia 22908-0736, USA.
Derewenda
Urszula
U
Devedjiev
Yancho
Y
Dauter
Zbigniew
Z
Derewenda
Zygmunt S
ZS
D000818:Animals; D001921:Brain / Q000002:abnormalities; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology; D003460:Crystallization; D018360:Crystallography, X-Ray; D000089742:Doublecortin-Like Kinases; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008958:Models, Molecular; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology; D010215:Papio; D010446:Peptide Fragments / Q000737:chemistry; D017346:Protein Serine-Threonine Kinases; D012174:Retinitis Pigmentosa / Q000201:enzymology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D047908:Intracellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; C117347:DCLK1 protein, human; D000089742:Doublecortin-Like Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases