CK2-mediated phosphorylation of a type II regulatory subunit of cAMP-dependent protein kinase from the mollusk Mytilus galloprovincialis

Two isoforms of regulatory (R) subunit of cAMP-dependent protein kinase (PKA), named Rmyt1 and Rmyt2, were identified so far in the sea mussel Mytilus galloprovincialis. Out of them, only Rmyt2 was phosphorylated in vitro by casein kinase 2 (CK2) using GTP as phosphate donor. CK2 catalytic subunit (CK2α) itself was sufficient to phosphorylate Rmyt2, but phosphorylation was enhanced by the presence of the regulatory subunit CK2β. Even in the absence of CK2, Rmyt2 was phosphorylated to a certain extent when it was incubated with GTP. This basal phosphorylation was partially abolished by the known inhibitors apigenin and emodin, which suggests the presence of a residual amount of endogenous CK2 in the preparation of purified R subunit. CK2-mediated phosphorylation significantly decreases the ability of Rmyt2 to inhibit PKA catalytic (C) subunit activity in the absence of cAMP. On the other hand, the sequence of several peptides obtained from the tryptic digestion of Rmyt2 showed that mussel protein contains the signature sequence common to all PKA family members, within the “phosphate binding cassette” (PBC) A and B. Moreover, the degree of identity between the sequences of peptides from Rmyt2, as a whole, and those from type II R subunits was 68–75%, but the global identity percentage with type I R subunits was only about 30%, so that Rmyt2 can be classified as a type II R subunit.