The Affinity of Calmodulin-Affigel for Inositol Triphosphate Kinase From Bovine Brain.
10.12701/yujm.1990.7.1.39
- Author:
Sung Woo LIM
;
Jung Hye KIM
- Publication Type:Original Article
- MeSH:
Adenosine Triphosphate;
Adenylyl Cyclases;
Brain*;
Calmodulin;
Cholic Acid;
Chromatography;
Cyclic AMP;
Deoxycholic Acid;
Detergents;
Egtazic Acid;
Electrophoresis;
Hydrolysis;
Inositol*;
Membranes;
Metabolism;
Occupations;
Phosphotransferases*;
Polyethylene Glycols;
Polysorbates;
Second Messenger Systems;
Toes;
Type C Phospholipases
- From:Yeungnam University Journal of Medicine
1990;7(1):39-50
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
The one event on signaling mechanism is the cleavage by adenyl cyclase of ATP into second messenger, cyclic AMP. The other transfer system of inositol metabolism, it is widely recognized that hydrolysis of the minor membrane lipid phosphoinositide bisphosphate (PIP₂) initiated by occupation of certain receptors and catalyzed by phospholipase C, lead to toe generation of the two intracellular messengers, inositol triphosphate (IP₃) and diacylglycerol (DG). IP₃ is converted to inositol tetrakisphosphate (IP₄) by IP₃ kinase. In the present study, it is that purification of calmodulin is used by phenyl-Sepharose CL-4B chromatography, it's molecular weigh, 17,000 in SDS-polyacrylamide gel electrophoresis. In order to observe the affinity between calmodulin (CaM)-Affigel 15 and IP₃ kinase, and isolated IP₃ kinase, was applied in CaM-Affigel with Ca²⁺ equilibrium buffer and EGTA equilibrium buffer. We compared with binding and elution effect of IP₃ kinase in several condition of buffer. In affinity of binding, Ca²⁺ equilibrium buffer was in the most proper condition, and elution, CaM/Ca²⁺buffer (CE 1 10.36, CE2 12.76pM/min/mg of protein) was effected much more than EGTA buffer (E2 1.48, E 2.43pM/min/mg of protein), but CaM/Ca²⁺stimulate the activity of IP₃ kinase. And then, several detergents such as sodium deoxycholate, tween 20, cholic acid, polyethylene glycol, chaps were applied. The 0.2% chaps buffer (E2 23.19, E3 8.05pnM/min/mg of protein) was the most effective in elution of IP3 kinase.
