Many small G proteins, like K-Ras4B and Rnd3, possess flexible C-terminal tails with hydrophobic farnesylation and methylation, which mediate their interaction with specific membrane microdomains and distinct chaperon proteins. Moreover, it has been found that additional phosphate modifications on the tails are involved in the regulation of the abovementioned biomolecular interactions. The signaling activities of K-Ras4B and Rnd3 are highly associated with their cellular localization. However, the molecular mechanism by which farnesylation and phosphorylation co-regulate the interaction networks of K-Ras4B or Rnd3 remains elusive. Protein chemical total synthesis and semi-synthesis provide very powerful tools for generating chemical structure-defined proteins with different PTMs, which can serve as useful probes or potential modulators for PTM-involved biological processes [1]. Thus, we respectively synthesized these two proteins with farnesyl, phosphate and methyl modifications. By using these homogeneous synthetic proteins with different PTM patterns, we reveal that phosphorylation does not fully inhibit the membrane association of K-Ras4B but rather weakens its interaction with membranes in a membrane fluidity-dependent manner [2]. In addition, we demonstrate that remote phosphate modifications mediate the multivalent interaction between Rnd3 and chaperon protein 14-3-3x through an additive mechanism, whereas adjacent phosphate and farnesyl modifications cooperatively contribute to the interaction by following an induced fit mechanism [3]. These mechanistic insights provide an basis for further developing modulators of K-Ras4B-membrane interaction or Rnd3-14-3-3x interaction.