title: Paramecium has two regulatory subunits of cyclic AMP-dependent protein kinase, one unique to cilia.
abstract: The subunit composition and intracellular location of the two forms of cAMP-dependent protein kinase of Paramecium cilia were determined using antibodies against the 40-kDa catalytic (C) and 44-kDa regulatory (R44) subunits of the 70-kDa cAMP-dependent protein kinase purified from deciliated cell bodies. Both C and R44 were present in soluble and particulate fractions of cilia and deciliated cells. Crude cilia and a soluble ciliary extract contained a 48-kDa protein (R48) weakly recognized by one of several monoclonal antibodies against R44, but not recognized by an anti-R44 polyclonal serum. Gel-filtration chromatography of a soluble ciliary extract resolved a 220-kDa form containing C and R48 and a 70-kDa form containing C and R44. In the large enzyme, R48 was the only protein to be autophosphorylated under conditions that allow autophosphorylation of R44. The subunits of the large enzyme subsequently were purified to homogeneity by cAMP-agarose chromatography. Both C and R48 were retained by the column and eluted with I M NaCl; no other proteins were purified in this step. These results confirm that the ciliary cAMP-dependent protein kinases have indistinguishable C subunits, but different R subunits. The small ciliary enzyme, like the cell-body enzyme, contains R44, whereas R48 is the R subunit of the large enzyme.
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison 53706-1569, USA.
authors.forename: M
authors.initials: M
authors.lastname: Hochstrasser
authors.forename: G L
authors.initials: GL
authors.lastname: Carlson
authors.forename: C E
authors.initials: CE
authors.lastname: Walczak
authors.forename: D L
authors.initials: DL
authors.lastname: Nelson
mesh_terms: D000818:Animals; D002847:Chromatography, Agarose; D002850:Chromatography, Gel; D002923:Cilia / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D015151:Immunoblotting; D016805:Paramecium tetraurelia / Q000201:enzymology*; D010766:Phosphorylation; D012045:Regulatory Sequences, Nucleic Acid
chemical_list: D017868:Cyclic AMP-Dependent Protein Kinases