Thermoresponsive gels containing gold nanoparticles (AuNPs) were prepared using Pluronic®127 alone (F1) and with hydroxypropyl methylcellulose (F2) at ratios of 15% w/w and 15:1% w/w, respectively. AuNPs were evaluated for particle size, zeta-potential, polydispersity index (PDI), morphology and XRD pattern. AuNP-containing thermoresponsive gels were investigated for their gelation temperature, gel strength, bio-adhesive force, viscosity, drug content, in vitro release and ex-vivo permeation, in addition to in vitro antibacterial activity against bacteria found in burn infections, Staphylococcus aureus. In vivo burn healing and antibacterial activities were also investigated and compared with those of a commercial product using burn-induced infected wounds in mice. Spherical AuNPs sized 28.9–37.65 nm displayed a surface plasmon resonance band at 522 nm, a PDI of 0.461, and a zeta potential of 34.8 mV with a negative surface charge. F1 and F2 showed gelation temperatures of 37.2 °C and 32.3 °C, bio-adhesive forces of 2.45 ± 0.52 and 4.76 ± 0.84 dyne/cm2, viscosities of 10,165 ± 1.54 and 14,213 ± 2.31 cP, and gel strengths between 7.4 and 10.3 sec, respectively. The in vitro release values of F1 and F2 were 100% and 98.03% after 6 h, with permeation flux values of (J1) 0.2974 ± 2.85 and (J2) 0.2649 ± 1.43 (µg/cm2·h), respectively. The formulations showed antibacterial activity with the highest values for wound healing properties, as shown in vivo and by histopathological studies. This study demonstrates that a smart AuNPs thermoresponsive gel was successful as an antibacterial and wound healing transdermal drug delivery system.