Role of the beta subunit of casein kinase-2 on the stability and specificity of the recombinant reconstituted holoenzyme.
Recombinant human alpha subunit from casein kinase-2 (CK-2) was subjected, either alone or in combination with recombinant human beta subunit, to high temperature, tryptic digestion and urea treatment. In all three cases, it was shown that the presence of the beta subunit could drastically reduce the loss of kinase activity, strongly suggesting a protective function for the beta subunit. Assaying different peptides for specificity toward the recombinant alpha subunit and the recombinant reconstituted enzyme, showed that the presence of the beta subunit could modify the specificity of the catalytic alpha subunit. Therefore, a dual function for the beta subunit is proposed which confers both specificity and stability to the catalytic alpha subunit within the CK-2 holoenzyme complex. The peptide DLEPDEELEDNPNQSDL, reproducing the highly acidic amino acid 55-71 segment of the human beta subunit, counteracts the stimulatory effect of the beta subunit on the alpha subunit activity and partially substitutes the beta subunit in conferring thermal stability to the alpha subunit. No such effect is induced by the peptide MSSSEEVSW, reproducing the N-terminal segment of the beta subunit including the autophosphorylation site. It is suggested that the acidic domain of the beta subunit, encompassing residues 55-71, plays a role in the interactions between the beta and alpha subunits.
Meggio
F
F
Dipartimento di Chimica Biologica, Università di Padova, Italy.
Boldyreff
B
B
Marin
O
O
Pinna
L A
LA
Issinger
O G
OG
D000595:Amino Acid Sequence; D000818:Animals; D047388:Casein Kinases; D004795:Enzyme Stability; D006358:Hot Temperature; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D046911:Macromolecular Substances; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014508:Urea / Q000494:pharmacology
D046911:Macromolecular Substances; D011994:Recombinant Proteins; D014508:Urea; D011494:Protein Kinases; D047388:Casein Kinases; D008274:Magnesium