To enhance the mechanical properties and moisture barrier performance of cassava starch (CS) films, this study employed peanut protein isolate (PPI) and gelatin (GL) as protein modifiers to prepare composite films with CS. The effects of protein sources and addition ratio (0:10, 1:9, 2:8, 3:7, 4:6, and 5:5) on the properties and microstructure of the composite films were systematically investigated. The results showed that both proteins demonstrated good compatibility with CS, effectively enhancing the mechanical properties, water barrier performance, and microstructure density of the films. In the PPI/CS composite film, the optimal mass ratio was 3:7. The moisture content, water solubility, water absorption, moisture absorption, and water vapor permeability of the film were reduced by 28.32%, 19.39%, 14.79%, 27.87%, and 11.98%, respectively, compared to those of the pure CS film. The elongation at break also increased from 53.39% to 82.16%. In the GL/CS composite film, the addition of GL primarily improved the tensile strength of the film. When the mass ratio of GL/CS was 4:6, the tensile strength increased from 2.36 to 4.44 MPa, and the water barrier properties were significantly improved (P<0.05). Scanning electron microscopy and X-ray diffraction analyses indicated that protein addition optimized the surface morphology and modified the crystalline structure of the films, whereas Fourier-transform infrared spectroscopy confirmed the formation of hydrogen bonds between protein and starch molecules. In summary, PPI/CS composite films are more appropriate for short-term packaging requiring high flexibility and moisture barrier performance, whereas GL/CS composite films are more suitable for food packaging applications requiring greater structural stability. Both composites demonstrate significant potential for application in the field of biodegradable packaging materials.