The heterotrimeric G protein subunits Gαq and Gβ1 have lysophospholipase D activity

In a previous study we purified a novel lysoPLD (lysophospholipase D) which converts LPC (lysophosphatidylcholine) into a bioactive phospholipid, LPA (lysophosphatidic acid), from the rat brain. In the present study, we identified the purified 42 and 35 kDa proteins as the heterotrimeric G protein subunits Gαq and Gβ1 respectively. When FLAG-tagged Gαq or Gβ1 was expressed in cells and purified, significant lysoPLD activity was observed in the microsomal fractions. Levels of the hydrolysed product choline increased over time, and the Mg2+ dependency and substrate specificity of Gαq were similar to those of lysoPLD purified from the rat brain. Mutation of Gαq at amino acids Lys52, Thr186 or Asp205, residues that are predicted to interact with nucleotide phosphates or catalytic Mg2+, dramatically reduced lysoPLD activity. GTP does not compete with LPC for the lysoPLD activity, indicating that these substrate-binding sites are not identical. Whereas the enzyme activity of highly purified FLAG-tagged Gαq overexpressed in COS-7 cells was ~4 nmol/min per mg, the activity from Neuro2A cells was 137.4 nmol/min per mg. The calculated Km and Vmax values for lysoPAF (1-O-hexadecyl-sn-glycero-3-phosphocholine) obtained from Neuro2A cells were 21 μM and 0.16 μmol/min per mg respectively, similar to the enzyme purified from the rat brain. These results reveal a new function for Gαq and Gβ1 as an enzyme with lysoPLD activity. Tag-purified Gα11 also exhibited a high lysoPLD activity, but Gαi and Gαs did not. The lysoPLD activity of the Gα subunit is strictly dependent on its subfamily and might be important for cellular responses. However, treatment of Hepa-1 cells with Gαq and Gα11 siRNAs (small interfering RNAs) did not change lysoPLD activity in the microsomal fraction. Clarification of the physiological relevance of lysoPLD activity of these proteins will need further studies.