Tyrosine versus serine/threonine phosphorylation by protein kinase casein kinase-2 - A study with peptide substrates derived from immunophilin Fpr3

Protein kinase casein kinase-2 (CK2) is a spontaneously active, ubiquitous, and pleiotropic enzyme that phosphorylates seryl/threonyl residues specifi ed by multiple negatively charged side chains, the one at position n + 3 be ing of crucial importance (minimum consensus S/T-x-x-E/D/S(P)/T(P). Recentl y CK2 has been reported to catalyze phosphorylation of the yeast nucleolar immunophilin Fpr3 at a tyrosyl residue (Tyr(184)) fulfilling the consensus sequence of Ser/Thr substrates (Wilson, L.K, Dhillon, N., Thorner, J., and Martin, G.S. (1997) J. Biol Chem. 272, 12961-12967). Here we show that, by contrast to other tyrosyl peptides fulfilling the consensus sequence for CK 2, a peptide reproducing the sequence around Fpr3 Tyr(184) (DEDADIY(184)DEE DYDL) is phosphorylated by CK2, albeit with much higher K-m (384 versus 4.3 mu M) and lower V-max (8.4 versus 1,132 nmol.min(-1). mg(-1)) than its der ivative with Tyr(184) replaced by serine. The replacement of Asp at positio n n + 1 with alanine and, to a lesser extent, of Ile at n - 1 with Asp are especially detrimental to tyrosine phosphorylation as compared with serine phosphorylation, which is actually stimulated by the ne to Asp modification . In contrast the replacement of Glu at n + 3 with alanine almost suppresse s serine phosphorylation but not tyrosine phosphorylation. It can be conclu ded that CK2 is capable to phosphorylate, under special circumstances, tyro syl residues, which are specified by structural features partially differen t from those that optimize Ser/Thr phosphorylation.