Identification of structural elements involved in the interaction of simian virus 40 small tumor antigen with protein phosphatase 2A.
SV40 small tumor antigen (small-t) was used as a model to identify structural elements involved in the interactions between regulatory proteins and protein phosphatase 2A (PP2A). Using mutant proteins and synthetic peptides, we identified a small domain within small-t that is a major site for interaction with the dimeric form of PP2A. A series of small-t truncation mutants identified a region surrounding the first of two conserved cysteine clusters that was critical for interaction with PP2A. These mutants also identified additional regions of small-t that contribute to high affinity interaction. Deletion of residues 110-119, which encompass the first cysteine cluster, resulted in a protein that failed to bind to PP2A. Synthetic peptides that contained residues 105-122 of small-t blocked binding of small-t to PP2A. These peptides also inhibited the phosphatase activity of PP2A in a manner analogous to full-length small-t. The active small-t peptides adopt a beta-strand structure that was essential for high affinity interaction with the PP2A dimer. Based on circular dichroism measurements, the same cysteine cluster-containing peptides that bind to PP2A also interact with zinc. Interaction with zinc required the conserved cysteines but was not required for interaction with PP2A.
Mateer
S C
SC
Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, Texas 75235-9041, USA.
Fedorov
S A
SA
Mumby
M C
MC
D000595:Amino Acid Sequence; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002942:Circular Dichroism; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000187:drug effects / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D054648:Protein Phosphatase 2; D017386:Sequence Homology, Amino Acid; D013539:Simian virus 40 / Q000276:immunology*; D015032:Zinc / Q000494:pharmacology
D000952:Antigens, Polyomavirus Transforming; D010446:Peptide Fragments; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D015032:Zinc