To further understand the high-pressure treatment (HPT) induced modifications to meat proteins, extracted myosin (0.6 M KCl, 20 mM K2HPO4/KH2PO4, pH 6.5) was subjected to 100–300 MPa HPT (9 min, 25 °C). Focusing on the micro-perspective aspects, Raman spectra, dynamic scanning fluorimetry (DSF), scanning electron microscope (SEM) was applied to either myosin solutions or the corresponding thermal gels. Results revealed that micro-environmental properties almost remain intact following HPT, except for the more buried tyrosine residues and the up-shifting of wavenumber near 1460 cm−1; more discrepancies appeared subsequent to formation of myosin thermally-induced gels, especially the amide Ⅰ, Ⅲ bands and the OH-stretching vibrations. Thermal stability of 200, 300 MPa HPT-treated myosin was significantly improved (P < 0.05) based on the DSF. Visualization and fractal analysis of SEM images revealed that myosin, following 200, 300 MPa HPT formed thicker stranded gels, exhibiting lower complexity and larger cavities. These results extend the understanding of HPT-induced protein modifications relating to gelation functionalities.