Purification and characterization of casein kinase II (CKII) from delta cka1 delta cka2 Saccharomyces cerevisiae rescued by Drosophila CKII subunits. The free catalytic subunit of casein kinase II is not toxic in vivo.
Casein kinase II (CKII) is composed of a catalytic (alpha) and a regulatory (beta) subunit which unite to form an alpha 2 beta 2 holoenzyme. Saccharomyces cerevisiae CKII consists of two distinct catalytic (Sc alpha and Sc alpha') and regulatory (Sc beta and Sc beta') subunits. Simultaneous disruption of the CKA1 and CKA2 genes (encoding the alpha and alpha' subunits, respectively) is lethal. Such double disruptions can be rescued by GAL1, 10-induced expression of the Drosophila alpha and beta subunits (Dm alpha+beta) together or by GAL10-induced expression of the Drosophila alpha subunit (Dm alpha) alone (Padmanabha, R., Chen-Wu, J. L.-P., Hanna, D. E., and Glover, C. V. C. (1990) Mol. Cell. Biol. 10, 4089-4099). Here we report quantitation, purification, and characterization of casein kinase II activity from such rescued strains. Casein kinase II activity from a strain rescued by Dm alpha alone purifies as a free, catalytically active alpha subunit monomer, whereas that from a strain rescued by Dm alpha/beta purifies as a mixture of tetrameric holoenzyme and monomeric alpha subunit. Interestingly, neither Sc beta nor Sc beta' is present at detectable levels in the enzyme obtained from either strain, raising the possibility that rescue by Dm alpha alone may be mediated via the free, monomeric catalytic subunit. Overexpression of total casein kinase II activity from 6- to 18-fold is not toxic and indeed has no overt phenotypic consequences. Production of large amounts of free catalytic subunit also appears to be without effect, even though free catalytic subunit is normally undetectable in S. cerevisiae.
Bidwai
A P
AP
Department of Biochemistry, University of Georgia, Athens 30602.
Hanna
D E
DE
Glover
C V
CV
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D047390:Casein Kinase II; D002384:Catalysis; D002853:Chromatography, Liquid; D004330:Drosophila / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010957:Plasmids; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases