Purification of brain-type creatine kinase (B-CK) from several tissues of the chicken: B-CK subspecies.
A method for the purification of brain-type creatine kinase (B-CK) from several tissues of the chicken, e.g., brain, retina, gizzard and heart was developed involving (1) an affinity chromatography step on Sepharose Blue from which B-CK was specifically eluted by ADP and (2) a subsequent anion exchange chromatography step on a fast protein liquid chromatography Mono-Q column. Two distinct peaks with B-CK activity, both purified to greater than or equal to 99% homogeneity and displaying specific enzyme activities of 300-400 mumol CP/min/mg 1t pH 7.0 and 25 degrees C, were eluted by a salt gradient at a plateau of 150 mmol/l NaCl. The ratio of the two B-CK peaks varied in a tissue-dependent manner, indicating that in chicken the dimerization of native BB-CK from the two major B-CK subunit species is tissue-specific and nonrandom in neural tissues. The fast, efficient and convenient method for the purification of B-CK at small or large scale, operating at yields of 50-70%, makes the purification of this rather labile enzyme from small amounts of tissues possible and greatly facilitates the subsequent characterization of both major and minor dimeric BB-CK subspecies present in these different tissues.
Quest
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AF
Institute of Cell Biology, ETH-Hönggerberg, Zürich, Switzerland.
Eppenberger
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HM
Wallimann
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T
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002645:Chickens; D003402:Creatine Kinase / Q000302:isolation & purification* / Q000378:metabolism; D005895:Gizzard, Avian / Q000201:enzymology; D007527:Isoenzymes; D007700:Kinetics; D009130:Muscle, Smooth / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology; D009928:Organ Specificity; D012160:Retina / Q000201:enzymology
D007527:Isoenzymes; D003402:Creatine Kinase