Objective: Impaired glycemic control is a common comorbidity of obesity. E4orf1(E4), an adenovirus-derived protein, reduces the activity of insulin receptor substrate (IRS), yet activates Akt and promotes the membrane translocation of GLUT4, resulting in better glycemic control in mice. To develop a clinically suitable delivery system, here we constructed and tested liposome nanoparticles (NP), to deliver E4 to preadipocytes. Methods: Glutathione-S-transferase (GST)-tagged E4 was encapsulated in Rhodamine-phosphatidylethanolamine (PE)-tagged soy-phosphatidylcholine-NP. The NP were characterized. Preadipocytes were treated with free E4, E4 containing NP (E4 NP) or E4-free NP (void NP). Results: For void and E4 NP, the average size was ~150 and 130 nm, PDI was ~0.25 and 0.27, and Zeta potential was −23 and −25, respectively. The average encapsulation efficiency (EE) was ~50%. Cells treated with E4 showed maximum GST expression and Rhodamine signals at 24 h. The presence of E4 in cells was confirmed at 24, 48, and 72 h. At 72 h after exposure, E4 NP significantly decreased pTyr-IRS, yet increased pAkt protein abundance, membrane translocation of GLUT4, and glucose uptake, compared with cells treated with void NP. Free E4 (without NP) had no effect. Conclusions: NP-mediated delivery of E4 promotes glucose uptake in preadipocytes. The next step is to test the efficacy of this clinically compatible delivery approach in vivo.