Two novel β-galactosidases from Aeromonas caviae with potential industrial applications in milk and catalytic mechanism analysis using molecular docking.

Lactose intolerance has been a significant global health concern, as it is caused by the absence of lactase, leading to the inability of the human body to absorb lactose. This study investigated two genes encoding maltose O-acetyltransferase with β-galactosidase activity from Aeromonas caviae to evaluate their potential application value for lactose degradation in bovine and human milk. The two novel β-galactosidases (AcGal25: 22.0 kDa and AcGal31: 21.3 kDa) were heterologously expressed and biochemically characterized. The optimal pH of both enzymes was 8.0, and the optimal temperature of AcGal25 and AcGal31 were 45 and 42 °C, respectively. Fe2+ and Mn2+ significantly promoted the activity of both enzymes. The two enzymes kept over 75 % activity after incubation for 30 days at 45 °C. HPLC results showed that lactose in bovine milk was completely hydrolyzed by AcGal31 when reacted for 6 h, and about 5 % lactose in human milk was left. The docking results showed that AcGal31 has a more vital lactose-binding ability than AcGal25. LYS129 and ARG165 are highly likely catalytic sites of AcGal31. AcGal31 demonstrated excellent commercial value in making lactose-free milk.