Overexpression, purification, and preliminary X-ray crystallographic analysis of human brain-type creatine kinase.
Creatine kinase (CK; E.C. 2.7.3.2) is an important enzyme that catalyzes the reversible transfer of a phosphoryl group from ATP to creatine in energy homeostasis. The brain-type cytosolic isoform of creatine kinase (BB-CK), which is found mainly in the brain and retina, is a key enzyme in brain energy metabolism, because high-energy phosphates are transferred through the creatine kinase/phosphocreatine shuttle system. The recombinant human BB-CK protein was overexpressed as a soluble form in Escherichia coli and crystallized at 22 degrees C using PEG 4000 as a precipitant. Native X-ray diffraction data were collected to 2.2 A resolution using synchrotron radiation. The crystals belonged to the tetragonal space group P43212, with cell parameters of a=b=97.963, c= 164.312 A, and alpha=beta=gamma=90 degrees. The asymmetric unit contained two molecules of CK, giving a crystal volume per protein mass (Vm) of 1.80 A3 Da-1 and a solvent content of 31.6%.
Bong
Seung Min
SM
Division of Biotechnology, College of Life Sciences, Korea University, Seoul 136-713, Korea.
Moon
Jin Ho
JH
Jang
Eun Hyuk
EH
Lee
Ki Seog
KS
Chi
Young Min
YM
D052276:Creatine Kinase, BB Form / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D011994:Recombinant Proteins; D052276:Creatine Kinase, BB Form