The tetratricopeptide repeat domain and a C-terminal region control the activity of Ser/Thr protein phosphatase 5.
Protein Ser/Thr phosphatase 5 is a 58-kDa protein containing a catalytic domain structurally related to the catalytic subunits of protein phosphatases 1, 2A, and 2B and an extended N-terminal domain with three tetratricopeptide repeats. The activity of this enzyme is stimulated 4-14-fold in vitro by polyunsaturated fatty acids and anionic phospholipids. The structural basis for lipid activation of protein phosphatase 5 was examined by limited proteolysis and site-directed mutagenesis. Trypsinolysis removed the tetratricopeptide repeat domain and increased activity to approximately half that of lipid-stimulated, full-length enzyme. Subtilisin removed the tetratricopeptide repeat domain and 10 residues from the C terminus, creating a catalytic fragment with activity that was equal to or greater than that of lipid-stimulated, full-length enzyme. Catalytic fragments generated by proteolysis were no longer stimulated by lipid, and degradation of the tetratricopeptide repeat domain was decreased by association with lipid. A truncated mutant missing 13 C-terminal residues was also insensitive to lipid and was as active as full-length, lipid-stimulated enzyme. These results suggest that the C-terminal and N-terminal domain act in a coordinated manner to suppress the activity of protein phosphatase 5 and mediate its activation by lipid. These regions may be targets for the regulation of protein phosphatase 5 activity in vivo.
Sinclair
C
C
Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907, USA.
Borchers
C
C
Parker
C
C
Tomer
K
K
Charbonneau
H
H
Rossie
S
S
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D004789:Enzyme Activation; D006868:Hydrolysis; D050356:Lipid Metabolism; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding
D009687:Nuclear Proteins; D010455:Peptides; D010749:Phosphoprotein Phosphatases; C089641:protein phosphatase 5