Characterization of Saccharomyces cerevisiae protein Ser/Thr phosphatase T1 and comparison to its mammalian homolog PP5
Department of Biochemistry, Purdue University, West Lafayette, IN 47907, USA
BMC Cell Biology 2003, 4:3 doi:10.1186/1471-2121-4-3Published: 28 March 2003
Protein Ser/Thr phosphatase 5 (PP5) and its Saccharomyces cerevisiae homolog protein phosphatase T1 (Ppt1p) each contain an N-terminal domain consisting of several tetratricopeptide repeats (TPRs) and a C-terminal catalytic domain that is related to the catalytic subunits of protein phosphatases 1 and 2A, and calcineurin. Analysis of yeast Ppt1p could provide important clues to the function of PP5 and its homologs, however it has not yet been characterized at the biochemical or cellular level.
The specific activity of recombinant Ppt1p toward the artificial substrates 32P-myelin basic protein (MBP) and 32P-casein was similar to that of PP5. Dephosphorylation of 32P-MBP, but not 32P-casein, was stimulated by unsaturated fatty acids and by arachidoyl coenzyme A. Limited proteolysis of Ppt1p removed the TPR domain and abrogated lipid stimulation. The remaining catalytic fragment exhibited a two-fold increase in activity toward 32P-MBP, but not 32P-casein. Removal of the C terminus increased Ppt1p activity toward both substrates two fold, but did not prevent further stimulation of activity toward 32P-MBP by lipid treatment. Ppt1p was localized throughout the cell including the nucleus. Levels of PPT1 mRNA and protein peaked in early log phase growth.
Many characteristics of Ppt1p are similar to those of PP5, including stimulation of phosphatase activity with some substrates by lipids, and peak expression during periods of rapid cell growth. Unlike PP5, however, proteolytic removal of the TPR domain or C-terminal truncation only modestly increased its activity. In addition, C-terminal truncation did not prevent further activation by lipid. This suggests that these regions play only a minor role in controlling its activity compared to PP5. Ppt1p is present in both the nucleus and cytoplasm, indicating that it may function in multiple compartments. The observation that Ppt1p is most highly expressed during early log phase growth suggests that this enzyme is involved in cell growth or its expression is controlled by metabolic or nutritional signals.