Investigation of a conserved tyrosine residue in a yeast casein kinase 1 protein kinase

Abstract

Yck1/2 are essential and redundant plasma membrane‐associated casein kinase 1 (CK1) proteins in budding yeast. In addition to sequence features common to protein kinases, CK1s share features unique to this subfamily. We identified the peptide HIPYRE as such a feature and investigated its role in the function of Yck2 (H233‐E238). CK1s from S. pombe were shown to exhibit dual specificity and to be affected by Tyr phosphorylation. We find that Yck2 is Tyr phosphorylated, and we investigated the possibility that Yck2 activity is regulated by phosphorylation of Y236. We generated mutant YCK2 alleles to investigate growth phenotypes and in vitro kinase activity. Cells expressing only Yck2 deleted for Y236 show defects in common with the low activity yckts mutant, and the mutant protein shows decreased activity on casein in vitro. Y236S and Y236F substitutions do not appear to impair Yck2 activity in vitro or in vivo, but the Y236E protein neither sustains normal growth of yeast nor phosphorylates casein at normal levels in vitro. We conclude that Y236 is important for activity, and that Yck2 activity could be negatively regulated by phosphorylation at this site. Work supported by NSF grant MCB‐0517204.