Open AccessRapid purification and characterization of protein kinase C from bovine retinal rod outer segments
Author(s) -
WOLBRING Gregor,
COOK Neil J.
Publication year - 1991
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1991.tb16320.x
Subject(s) - retinal , characterization (materials science) , chemistry , protein kinase c , microbiology and biotechnology , biology , biochemistry , kinase , materials science , nanotechnology
A rapid FPLC procedure for the purification of protein kinase C from bovine rod outer segments is described. The enzyme is essentially homogenous after purification and exhibits a molecular mass of approximately 85 kDa, as determined by SDS/PAGE. From its chromatographic behaviour on hydroxyapatite, and from Western‐blotting experiments using isoenzyme‐specific antibodies, we were able to identify the bovine rod outer segment protein kinase C as being of the α or type‐III form. The purified protein kinase C has a specific activity of 1066 nmol 32 P · min −1 · mg protein −1 , and shows a 30‐fold activation upon the addition of the effectors Ca 2+ , PtdSer and 1,2‐diacylglycerol. Arachidonic acid and linoleic acid were also found to enhance significantly the activity of the purified enzyme.