Abstract: Infrared spectroscopy was used to study the secondary structure of yeast proteins. One-dimensional and second-derivative infrared spectroscopy along with deconvolution infrared spectroscopy of amide Ⅲ bands at different temperature were measured. It was found that with increasing temperature,the infrared absorption intensity of the α-helix structure of yeast protein decreased,while the absorption intensities of β-turn,random coil and β-sheet structures increased. Two-dimensional infrared spectroscopy of yeast amide Ⅲ bands was used to determine the order of the absorption band intensity changes. Thermal instability of the β-sheet structure of yeast protein was further confirmed.