Identification of phosphorylation sites in the repetitive carboxyl-terminal domain of the mouse RNA polymerase II largest subunit.

Abstract

The largest subunit of eukaryotic RNA polymerase II contains a carboxyl-terminal domain (CTD) which is comprised of repetitive heptapeptides with a consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. We demonstrate here that the mouse CTD expressed in and purified from Escherichia coli can be phosphorylated in vitro by a p34cdc2/CDC28-containing CTD kinase from mouse ascites tumor cells. The product of this reaction, a phosphorylated form of the CTD, contains phosphoserine and phosphothreonine, but not phosphotyrosine. The same phosphoamino acid content is observed in the in vivo phosphorylated CTD from a mouse cell line. Synthetic peptides with naturally occurring non-consensus heptapeptide sequences can also be phosphorylated by CTD kinase in vitro. Phosphoamino acid analysis of these non-consensus heptapeptides together with direct sequencing of a phosphorylated heptapeptide reveals that serines (or threonines) at positions two and five are the sites phosphorylated by mouse CTD kinase. Thus, the -Ser(Thr)-Pro- motif common to p34cdc2/CDC28-containing protein kinases is the recognition site for mouse CTD kinase.

• Authors: J Zhang, J L Corden
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 266 Issue 4 Pg. 2290-6 (Feb 05 1991) ISSN: 0021-9258 [Print] United States
• PMID: 1899239 (Publication Type: Journal Article)

Chemical References

• Amino Acids
• Oligopeptides
• Threonine
• Protein Kinases
• carboxy-terminal domain kinase
• RNA Polymerase II

Topics

• Amino Acid Sequence
• Amino Acids (analysis, metabolism)
• Animals
• Carcinoma, Ehrlich Tumor
• Electrophoresis, Polyacrylamide Gel
• Mice
• Molecular Sequence Data
• Oligopeptides (metabolism)
• Phosphorylation
• Protein Kinases (metabolism)
• RNA Polymerase II (chemistry, metabolism)
• Substrate Specificity
• Threonine (metabolism)
• Tumor Cells, Cultured