Goat milk β-casein is more easily digested and absorbed by infants than bovine milk β-casein, and the principal reason for this difference is the diversity of their structure. Recently, many investigations have reported the structure of bovine milk β-casein, however, the structure of goat milk β-casein and the difference between the structure of goat milk β-casein and bovine milk β-casein still needs to be studied in detail. The information of protein secondary structure can be obtained by spectroscopy technique, Circular dichroism is a method to characterize the structure of protein in solution state by using different optical active chromophores to absorb circularly polarized light in the left and right planes, which can make the protein conformation closer to its physiological state, and has the advantages of being rapid, simple and sensitive to conformational changes; Fourier transformation infrared spectroscopy is a method to characterize the structure of protein in solid state by using different chemical bonds or functional groups in the process of the vibration, which has the advantages of fast scanning speed, high resolution, wide measuring wavelength range, and is not easily affected by the molecular size and external conditions of protein samples. Circular dichroism and Fourier transformation infrared spectroscopy have been widely used in the study of protein conformation, but these two spectroscopy techniques to analyze the structure of β-casein has been rarely reported. Thus, this study used Circular dichroism and Fourier transformation infrared spectroscopy to compare the structural characteristics of the goat milk β-casein and bovine milk β-casein, and the sulfhydryl content and solubility of the two proteins were analyzed by spectrophotometry. Circular dichroism showed that random coil was the main secondary structure of goat milk β-casein and bovine milk β-casein, but the content of random coil of goat milk β-casein (50.2%±0.16%) was significantly higher than bovine milk β-casein (43.8%±0.14%), the content of α-helix (2.7%±0.21%) and β-fold (15.3%±0.08%) in the ordered structure were significantly lower than bovine milk β-casein (4.3%±0.13%, 19.5%±0.12%), the content of β-turn was 31.8%±0.11%, 32.4%±0.09%, respectively and the difference was not significant; Fourier transformation infrared spectroscopy showed that the content of α-helix, β-fold, β-turn in the secondary structure of goat milk β-casein were lower than bovine milk β-casein by 18%~20%, 9%~10%, 0.6%~1%, respectively and the content of random coil was higher than bovine milk β-casein by 17%~19%. The functional properties of the two proteins showed that the surface sulfhydryl content of goat milk β-casein and bovine milk β-casein were basically consistent with 19~20 μmol·g-1, but the total sulfhydryl content of goat milk β-casein (28.35±0.13 μmol·g-1) was significantly lower than bovine milk β-casein (46.72±0.21 μmol·g-1); the isoelectric point of goat milk β-casein was similar to bovine milk β-casein (pH 4~5), and the solubility of goat milk β-casein was lower than bovine milk β-casein near the isoelectric point, but higher than bovine milk β-casein far from the isoelectric point. The results showed that compared with the bovine milk β-casein, the disorder and flexibility of the goat milk β-casein were higher, and the internal structure of micelle was softer and looser.