The objective of this paper is to explore the effects of high hydrostatic pressure (HHP) treatments on the structural and physical properties of nisin-soy-protein-isolate (SPI) films by scanning electron microscopy (SEM), Fourier transformed infrared (FTIR) spectroscopy, circular dichroism (CD) spectroscopy, UV-Vis spectrophotometry, texture analyzer and rotational rheometer. It was found that the nisin-SPI based films treated by high HHP had uniform and smooth appearance and compact internal structures. Moreover, HHP treatment significantly improved the barrier and mechanical properties of the composite films. FTIR and CD analyses suggested that HHP treatments resulted in partial unfolding of protein molecules, which were supposed to lead to the improved structural and physical characteristics of the nisin-SPI films. Rheological analysis showed that all film-forming suspensions exhibited pseudo-plastic behavior, and HHP treatment significantly decreased the viscosity of the film-forming suspensions. Such effects were considered to contribute to finer structure and improved properties of the HHP films. The current results indicated that the structural transformation of SPI aggregates induced by HHP treatments introduced the changes in structural and physical properties of the films.