Sites important for PLCbeta2 activation by the G protein betagamma subunit map to the sides of the beta propeller structure

J Biol Chem. 1998 Oct 23;273(43):28298-304. doi: 10.1074/jbc.273.43.28298.

Abstract

The betagamma subunits of the heterotrimeric GTP-binding proteins (G proteins) that couple heptahelical, plasma membrane-bound receptors to intracellular effector enzymes or ion channels directly regulate several types of effectors, including phospholipase Cbeta and adenylyl cyclase. The beta subunit is made up of two structurally different regions: an N-terminal alpha helix followed by a toroidal structure made up of 7 blades, each of which is a twisted beta sheet composed of four anti-parallel beta strands (Wall, M. A., Coleman, D. E., Lee, E., Iñiguez-Lluhi, J. A., Posner, B. A., Gilman, A. G., and Sprang, S. R. (1995) Cell 83, 1047-1058; Lambright, D. G., Sondek, J., Bohm, A., Skiba, N. P., Hamm, H. E., and Sigler, P. B. (1996) Nature 379, 311-319). We have previously shown that sites for activation of PLCbeta2, PLCbeta3, and adenylyl cyclase II overlap on the "top" surface of the propeller, where Galpha also binds (Li, Y., Sternweis, P. M., Charnecki, S., Smith, T. F., Gilman, A. G., Neer, E. J., and Kozasa, T. (1998) J. Biol. Chem. 273, 16265-16272). The present study was undertaken to identify the regions on the side of the torus that might be important for effector interactions. We made mutations in each of the outer beta strands of the G protein beta1 propeller, as well as mutations in the loops that connect the outer strands to the adjacent beta strands. Our results suggest that activation of PLCbeta2 involves residues in the outer strands of blades 2, 6, and 7 of the propeller. We tested three of the mutations that most severely affected PLCbeta2 activity against two forms of adenylyl cyclase (ACI and ACII). Both inhibition of ACI and activation of ACII were unaffected by these mutations, suggesting that if ACI and ACII contact the outer strands, the sites of contact are different from those for PLCbeta2. We propose that distinct sets of contacts along the sides of the propeller will define the specificity of the interaction of betagamma with effectors.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenylyl Cyclases / metabolism
  • Animals
  • Binding Sites
  • Enzyme Activation
  • GTP-Binding Proteins / genetics
  • GTP-Binding Proteins / metabolism*
  • Humans
  • Isoenzymes / metabolism*
  • Models, Molecular
  • Mutagenesis
  • Phospholipase C beta
  • Protein Binding
  • Protein Conformation
  • Rats
  • Recombinant Proteins / metabolism
  • Type C Phospholipases / metabolism*

Substances

  • Isoenzymes
  • Recombinant Proteins
  • Type C Phospholipases
  • PLCB1 protein, human
  • Phospholipase C beta
  • Plcb1 protein, rat
  • GTP-Binding Proteins
  • Adenylyl Cyclases