Three warm-season genotypes of sorghum grain (Sorghum bicolor) have been developed. The objectives of this study were: (1) to systematically investigate the heat-related processing (HRP)-induced changes/alteration on: (a) detailed chemical and nutrient profiles, (b) protein subfractions, (c) energy values, (d) rumen degradation, (e) intestinal digestibility of rumen undegraded protein; and (2) to use vibrational non-invasive ATR-vFt/IR molecular spectroscopy to determine protein-related molecular structures in the warm-season genotypes of sorghum grain. The warm-season sorghum seeds were either maintained in their raw state (as control) or treated with the HRP using the Lindberg/Blue M™ for 80 min at 121 °C (the condition was chosen based on previous study). Compared with the control, the HRP treatment altered chemical and nutrient profiles and utilization and availability of warm-season sorghum grain. The HRP treatment increased metabolizable energy and net energy values. The HRP treatment also altered the protein molecular structure profile. Furthermore, it increased the ratio of Amide I and Amide II height and decreased the ratio of α-helix to β-sheet. In summary, the HRP treatment altered chemical nutrient profiles without negatively affect the nutritive value and digestion of warm-season sorghum grain. The molecular spectral profiles could be used as a potential predictor for the warm-season sorghum grain utilization and digestion.