Heterotrimeric G-proteins' activation on the intracellular side of the cell membrane is initiated by stimulation of the G-Protein Coupled Receptors (GPCRs) extra-cellular part. This two-step activation mechanism includes (1) an exchange between GDP and GTP molecules in the Gα subunit and (2) a dissociation of the whole Gαβγ complex into two membrane-anchored blocks, namely the isolated Gα and Gβγ subunits. Although X-ray data are available for both inactive Gαβγ:GDP and active Gα:GTP complexes, intermediate steps involved in the molecular mechanism of the dissociation have not yet been addressed at the molecular level. In this study, we first built a membrane-anchored intermediate Giαβγ:GTP complex. This model was then equilibrated by molecular dynamics simulations before the Targeted Molecular Dynamics (TMD) technique was used to force the Gα subunit to evolve from its inactive (GDP-bound) to its active (GTP-bound) conformations, as described by available X-ray data. The TMD constraint was applied only to the Gα subunit so that the resulting global rearrangements acting on the whole Gαβγ:GTP heterotrimer could be analyzed. We showed how these mainly local conformational changes of Gα could initiate large domain:domain motions of the whole complex, the Gβγ behaving as an almost quasi-rigid block. This separation of the two Gα:GTP and Gβγ subunits required the loss of several interactions at the Gα:Gβγ interface that were reported. This study provided an atomistic view of the crucial intermediate step of the G-proteins activation, e.g., the dissociation, that could hardly be elucidated by the experiment.