Effects of Module Truncation of a New Alginate Lyase VxAly7C from Marine Vibrio xiamenensis QY104 on Biochemical Characteristics and Product Distribution.

Abstract

Alginate lyase has received extensive attention as an important tool for oligosaccharide preparation, pharmaceutical production, and energy biotransformation. Noncatalytic module carbohydrate-binding modules (CBM) have a major impact on the function of alginate lyases. Although the effects of two different families of CBMs on enzyme characteristics have been reported, the effect of two combined CBM32s on enzyme function has not been elucidated. Herein, we cloned and expressed a new multimodular alginate lyase, VxAly7C, from Vibrio xiamenensis QY104, consisting of two CBM32s at N-terminus and a polysaccharide lyase family 7 (PL7) at C-terminus. To explore the function of CBM32s in VxAly7C, full-length (VxAly7C-FL) and two truncated mutants, VxAly7C-TM1 (with the first CBM32 deleted) and VxAly7C-TM2 (with both CBM32s deleted), were characterized. The catalytic efficiency of recombinant VxAly7C-TM2 was 1.82 and 4.25 times higher than that of VxAly7C-TM1 and VxAly7C-FL, respectively, indicating that CBM32s had an antagonistic effect. However, CBM32s improved the temperature stability, the adaptability in an alkaline environment, and the preference for polyG. Moreover, CBM32s contributed to the production of tri- and tetrasaccharides, significantly affecting the end-product distribution. This study advances the understanding of module function and provides a reference for broader enzymatic applications and further enzymatic improvement and assembly.