Crystallization and preliminary crystallographic studies of human TGF-beta type II receptor ligand-binding domain.
Three constructs (residues 15-136, 22-136 and 27-136) of the truncated extracellular domain of human transforming growth factor beta type II receptor (TBRII) were overexpressed in Escherichia coli. The constructs are referred to as TBRII(15-136), TBRII(22-136) and TBRII(27-136). The refolded receptors were purified using a combination of ion-exchange and size-exclusion chromatography. The purified receptors have an apparent molecular weight of 14 kDa as judged by size-exclusion chromatography. In the crystallization trials, TBRII(15-136) and TBRII(22-136) formed mostly crystal-like spheres but failed to produce data-quality crystals. TBRII(27-136) yielded large single crystals from hanging drops using the vapor-diffusion procedure with PEG 2000 or 4000 at pH 5.0. The crystals diffracted to 1.05 A [using the X9B beamline operated at lambda = 1.0092 A of the National Synchrotron Light Source (NSLS) at the Brookhaven National Laboratory] and belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 35.5, b = 40.7, c = 76.2 A. There was one molecule in the asymmetric unit, which corresponds to a solvent content of 42.1%.
Boesen
Christian C
CC
Structural Immunology Section, Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, Maryland 20852, USA.
Motyka
Shawn A
SA
Patamawenu
Apisit
A
Sun
Peter D
PD
D002845:Chromatography; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray / Q000379:methods*; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000737:chemistry*; D017356:Synchrotrons; D014961:X-Ray Diffraction
D008024:Ligands; D018125:Receptors, Transforming Growth Factor beta; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II