Anti-head and anti-tail antibodies against distinct epitopes in the catalytic subunit of protein kinase A - Use in the study of the kinase splitting membranal proteinase KSMP

Protein kinases share a considerable sequence homology in their catalytic core (residues 40-300 in PKA). Each core is flanked by ''head'' and ''tail'' segments at its amino- and carboxy-termini, which are different in the various kinases. These end segments may play an important role in creating the preferential affinity of each kinase for its physiological substrates or regulatory ligands. Here we describe three antipeptide antibodies (alpha P-1, alpha P-2, and alpha P-3) that specifically recognize the head and tail segments of the catalytic subunit (C) of PKA. (i) alpha P-1 (against (6)A-K-23) react with C when denatured but not when in its native structure; (ii) alpha P-2 (against K-319-I-335), bind to the site in C cleaved by the kinase splitting membranal proteinase (KSMP) and inhibit this cleavage of C; (iii) alpha P-3 (against S-338-F-350) react with C but not with the KSMP cleavage product C', useful for detecting a KSMP-like activity in different tissues and subcellular loci. The combined use of the antibodies described here provides a strict definition of C, and thus a high degree of fidelity in its biorecognition.