We report the effects of the substrate temperature on the surface morphology of Molybdenum trioxide (MoO3) thin films and the electrically detailed examination of Au/MoO3/n–GaAs MOS heterojunction structure with the best homogeneity. MoO3 thin film was deposited both on soda-lime silicate glass as a thin film and n-type and (100) oriented GaAs substrates using RF magnetron sputtering method at substrate temperatures of room temperature, 100 °C, 200 °C, and 300 °C. The surface morphology of the MoO3 thin films was investigated by utilizing atomic force microscopy (AFM) and scanning electron microscopy (SEM) measurements. AFM and SEM results have shown that MoO3 thin film with a substrate temperature of 200 °C has the lowest surface roughness and the homogeneity of the film structures significantly enhances with increasing substrate temperature up to 200 °C. An increment in the roughness of the thin-film structure was detected at higher temperatures than 200 °C due to the deterioration of homogeneity. Therefore, we primarily focused on the MoO3 thin films produced at the substrate temperature of 200 °C to examine the electrical properties of Au/MoO3/n–GaAs MOS heterojunction device. In order to determine the electrical properties, temperature-dependent measurements were performed in between 200 and 400 K by steps of 25 K. The fundamental electrical parameters such as saturation current (), ideality factor (), and barrier height () were calculated by analyzing the forward bias curves at different temperatures. The series resistance () values of the device were also determined using the plot of structure resistance () vs applied bias voltage (), Thermionic Emission Theory, and Cheung and Cheung methods. The value of Au/MoO3/n–GaAs MOS heterojunction device shows an abnormal behavior of up to 350 K, which is the critical temperature value and tends to increase with increasing temperature. Above the critical temperature value, it exhibits ideal behavior.