Differential Activity of the G Protein β5γ2 Subunit at Receptors and Effectors*

The G protein β5 subunit differs substantially in amino acid sequence from the other known β subunits suggesting that βγ dimers containing this protein may play specialized roles in cell signaling. To examine the functional properties of the β5 subunit, recombinant β5γ2 dimers were purified from baculovirus-infected Sf9 insect cells using a strategy based on two affinity tags (hexahistidine and FLAG) engineered into the N terminus of the γ2 subunit (γ2HF). The function of the pure β5γ2HF dimers was examined in three assays: activation of pure phospholipase C-β in lipid vesicles; activation of recombinant, type II adenylyl cyclase expressed in Sf9 cell membranes; and coupling of α subunits to the endothelin B (ETB) and M1 muscarinic receptors. In each case, the efficacy of the β5γ2HF dimer was compared with that of the β1γ2HF dimer, which has demonstrated activity in these assays. The β5γ2HF dimer activated phospholipase C-β with a potency and efficacy similar to that of β1γ2 or β1γ2HF; however, it was markedly less effective than the β1γ2HF or β1γ2 dimer in its ability to activate type II adenylyl cyclase (EC50 of ∼700 nm versus 25 nm). Both the β5γ2HF and the β1γ2HF dimers supported coupling of M1 muscarinic receptors to the Gq α subunit. The ETB receptor coupled effectively to both the Gi and Gq α subunits in the presence of the β1γ2HF dimer. In contrast, the β5γ2HF dimer only supported coupling of the Gq α subunits to the ETB receptor and did not support coupling of the Gi α subunit. These results suggest that the β5γ2HF dimer binds selectively to Gq α subunits and does not activate the same set of effectors as dimers containing the β1subunit. Overall, the data support a specialized role for the β5 subunit in cell signaling.

• Publication year

  1998

• Venue

  Journal of Biological Chemistry

• Publication date

  1998-12-18

• Fields of study

  Biology, Medicine, Chemistry

• Identifiers
  DOI 10.1074/JBC.273.51.34429PMID 9852110
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• The observed activities support a specialized role for the β5 subunit in selectively binding Gq alpha subunits and engaging a different effector repertoire than β1-containing dimers.

  Confidence 0.91

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• At the endothelin B receptor, β1γ2HF supported coupling to both Gi and Gq alpha subunits, whereas β5γ2HF supported only Gq coupling and not Gi coupling.

  Confidence 0.96

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β5γ2HF and β1γ2HF both supported coupling of the M1 muscarinic receptor to the Gq alpha subunit.

  Confidence 0.95

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β5γ2HF was much less effective than β1γ2 or β1γ2HF at activating type II adenylyl cyclase, with an EC50 of about 700 nM versus about 25 nM.

  Confidence 0.96

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β5γ2HF activated phospholipase C-β with potency and efficacy similar to β1γ2 and β1γ2HF.

  Confidence 0.97

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review

• endothelin b receptor

  receptor, protein

  A receptor used in coupling assays to test whether G protein dimers support signaling to different alpha subunits.

  Aliases: ETB receptor, ETB

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• gi alpha subunit

  protein subunit, protein

  A G protein alpha subunit associated with Gi-family signaling and tested for receptor coupling in the assays.

  Aliases: Gi α subunit, Gi alpha

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• gq alpha subunit

  protein subunit, protein

  A G protein alpha subunit associated with Gq-family signaling and tested for receptor coupling in the assays.

  Aliases: Gq α subunit, Gq alpha

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• m1 muscarinic receptor

  receptor, protein

  A muscarinic acetylcholine receptor subtype used to assess G protein coupling in the receptor assays.

  Aliases: M1 receptor, muscarinic M1 receptor

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• phospholipase c-β

  enzyme, effector

  A lipid-associated effector enzyme tested in vesicle-based activation assays with purified G protein dimers.

  Aliases: PLC-β, PLC beta

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• type ii adenylyl cyclase

  enzyme, effector

  A recombinant adenylyl cyclase isoform expressed in Sf9 cell membranes and used as an effector in the activation assay.

  Aliases: adenylyl cyclase II, AC II

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β1γ2hf dimer

  protein complex, recombinant protein

  A recombinant G protein β1-γ2 heterodimer with the same engineered γ2 tags, used as a comparison dimer in the assays.

  Aliases: β1γ2, β1γ2HF

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β5 subunit

  protein subunit, protein

  A G protein beta subunit with distinct sequence features relative to other known beta subunits, central to the paper's comparisons.

  Aliases: beta5 subunit, G protein β5 subunit

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review
• β5γ2hf dimer

  protein complex, recombinant protein

  A recombinant G protein β5-γ2 heterodimer purified from Sf9 insect cells using engineered hexahistidine and FLAG tags on the γ2 subunit.

  Aliases: β5γ2, β5γ2HF

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewB (s683577b42) review

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