Abstract: To achieve the enzymatic synthesis of γ-glutamyl-S-allyl-L-cysteine (GSAC), γ-glutamyltranspeptidase from Bacillus licheniformis (BlGGT) was obtained through fermentation followed by ammonium sulfate precipitation, and its enzymatic properties were characterized. Using L-glutamine (L-Gln) as the γ-glutamyl donor and S-allyl-L-cysteine (SAC) as the acceptor, the reaction conditions for BlGGT-catalyzed synthesis of GSAC were optimized via single-factor and orthogonal experiments. The results showed that the optimal temperature for both transpeptidation and hydrolytic activities of BlGGT was 55 ℃. The optimal pH for the transpeptidation reaction was 9.0, whereas that for the hydrolysis reaction was 7.0. The metal ions Zn²⁺ significantly enhanced both the transpeptidation and hydrolytic activities of BlGGT (P<0.05), Mg²⁺ and Ca²⁺ also significantly promoted transpeptidation activity (P<0.05), while Cu²⁺, Fe³⁺ and Mn²⁺ exhibited significant inhibitory effects on both activities (P<0.05). The optimal reaction conditions for GSAC synthesis were determined as follows: BlGGT concentration of 1 mg/mL, molar ratio of L-Gln to SAC of 1:3, reaction temperature of 60 ℃, pH10.0, and reaction time of 6 h. Under these optimized conditions, the conversion yield of GSAC reached 19.10%. The present study elucidated the enzymatic characteristics of BlGGT and established an efficient enzymatic process for GSAC synthesis, providing a theoretical foundation and technical support for the high-yield production of GSAC.