Protein phosphatase 2A holoenzyme assembly: identification of contacts between B-family regulatory and scaffolding A subunits. Academic Article uri icon

start page

  • 20750

end page

  • 20755


  • Protein serine/threonine phosphatase (PP) 2A is a ubiquitous enzyme with pleiotropic functions. Trimeric PP2A consists of a structural A subunit, a catalytic C subunit, and a variable regulatory subunit. Variable subunits (B, B', and B" families) dictate PP2A substrate specificity and subcellular localization. B-family subunits contain seven WD repeats predicted to fold into a beta-propeller structure. We carried out mutagenesis of Bgamma to identify domains important for association with A and C subunits in vivo. Several internal deletions in Bgamma abolished coimmunoprecipitation of A and C subunits expressed in COS-M6 cells. In contrast, small N- and C-terminal Bgamma deletions had no effect on incorporation into the PP2A heterotrimer. Thus, holoenzyme association of B-family subunits requires multiple, precisely aligned contacts within a core beta-propeller domain. Charge-reversal mutagenesis of Bgamma identified a cluster of conserved critical residues in Bgamma WD repeats 3 and 4. Acidic substitution of paired basic residues in Bgamma (RR165EE) abolished association with wild-type A and C subunits, while fostering incorporation of Bgamma into a PP2A heterotrimer containing an A subunit with an opposite charge-reversal mutation (EE100RR). Thus, binding of A and B subunits requires electrostatic interactions between conserved pairs of glutamates and arginines. By expressing complementary charge-reversal mutants in neuronal PC6-3 cells, we further show that holoenzyme incorporation protects Bgamma from rapid degradation by the ubiquitin/proteasome pathway.

date/time value

  • 2002

Digital Object Identifier (DOI)

  • 10.1074/jbc.M202992200

PubMed Identifier

  • 11929880


  • 277


  • 23


  • Amino Acid Sequence
  • Animals
  • Cell Line
  • DNA, Complementary
  • Dimerization
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Phosphoprotein Phosphatases
  • Precipitin Tests
  • Protein Conformation
  • Protein Phosphatase 2
  • Rats
  • Sequence Homology, Amino Acid