Dawei Ni, Zhaolin Huang, Shuqi Zhang, Yang Yang, Xiaoyong Liu, Wei Xu, Wenli Zhang, Wanmeng Mu
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Improving the activity of an inulosucrase by rational engineering for the efficient biosynthesis of low-molecular-weight inulin
Inulin, a widely recognized prebiotic, has diverse applications across various industrial sectors. Although inulin is primarily produced through plant extraction, there is growing interest in enzymatic synthesis as an alternative. The enzymatic production of inulin from sucrose, which yields polymers with degrees of polymerization similar to those of plant-derived inulin, shows potential as a viable replacement for traditional extraction methods. In this study, an inulosucrase from Neobacillus bataviensis was identified, demonstrating a non-processive mechanism specifically tailored for synthesizing inulin with polymerization degrees ranging from 3 to approximately 40. The enzyme exhibited optimal activity at pH 6.5 and 55 °C, efficiently producing inulin with a yield of 50.6%. Ca2+ can improve the activity and thermostability of this enzyme. To enhance catalytic total activity, site-directed and truncated mutagenesis techniques were applied, resulting in the identification of a mutant, T149S, displaying a significant 57% increase in catalytic total activity. Molecular dynamics simulations unveiled that the heightened flexibility observed in three surface regions positively influenced enzymatic activity. This study not only contributes to the theoretical foundation for inulosucrase engineering but also presents a potential avenue for the production of inulin.
期刊介绍:
Research papers must make a significant and original contribution to
microbiology and be of interest to a broad readership. The results of any
experimental approach that meets these objectives are welcome, particularly
biochemical, molecular genetic, physiological, and/or physical investigations into
microbial cells and their interactions with their environments, including their eukaryotic hosts.
Mini-reviews in areas of special topical interest and papers on medical microbiology, ecology and systematics, including description of novel taxa, are also published.
Theoretical papers and those that report on the analysis or ''mining'' of data are
acceptable in principle if new information, interpretations, or hypotheses
emerge.