Protein hydrolysates were prepared from an industrially defatted walnut meal (DWMPH) by enzymolysis employing Neutrase, Protamex, and Flavorzyme, respectively, with/without ultrasonic treatment. The effects of DWMPH supplementations on fermentation performance of lager yeast in high-gravity brewing were investigated. Results showed that ultrasonic-assisted enzymolysis simultaneous treatment (UAE) and ultrasonic pretreatment followed by enzymolysis (UPE) significantly increased degree of hydrolysis (DH) by 1.43 times and 0.71 times of traditional enzymolysis (TE) at least, respectively, Protamex treatment exhibited higher DH (13.3-32.8%) than Neutrase (9.2-25.3%) or Flavorzyme (11.8-28.7%). Compared with control, DWMPH supplementations prepared by UAE using Protamex (UAE-P), Neutrase (UAE-N), or Flavorzyme (UAE-F) significantly improved fermentation performance of lager yeast, especially for UAE-P with the highest major fractions of Mw < 1 kDa, increased wort fermentability and ethanol production by 15% and 17%, respectively, while UAE-F with the highest major fractions of Mw > 3 kDa obviously improved the foam stability of final beers. Furthermore, DWMPH supplementations significantly increased yeast growth and cell viability, promoted glycogen and trehalose accumulation, upregulated stress markers HSP12 and SSA3 expression in yeast cells, improved the formation of higher alcohols and esters, and increased the ratio of higher alcohol to ester indicating a better balanced taste of final beers.