Production of three oligosaccharide-metabolizing enzymes by reconstituted Pichia pastoris and its effect on honey bees
Year: 2025
Type: article
Abstract: Due to self-incompatibility, Camellia’s pollination efficiency is low. When honey bees are introduced for pollination, the raffinose oligosaccharides in the nectar induce bee poisoning. The purpose of this study is to reconstruct Pichia pastoris to express three key enzymes of galactose metabolism (GALK, GALT, GALE) in Andrena camellia to explore their effect on the detoxification ability of oligosaccharides in honeybees. The pPIC9K-GALK/GALT/GALE expression vector was constructed using plasmid recombination technology, which was then electrotransformed into Pichia pastoris GS115 and induced to produce the recombinant enzyme. Protein expression was verified by Western blot. In in vitro experiments, we fed honey bees sugar water supplemented with the recombinant enzymes. We then evaluated the metabolic efficiency of the enzymes in galactose, the survival rate, and the median survival of the bees. The results showed that the efficient expression of GALK, GALT, and GALE in P. pastoris was successfully achieved. In in vitro digestion experiments, the combination of three enzymes could significantly reduce the content of galactose (P<0.05), and we detected Leloir pathway metabolites (such as UDP-glucose). The feeding experiment showed that the median survival of the worker bees supplemented with three enzymes was 34.43% longer than that of the control group (P<0.05). GALK, GALT, and GALE expressed by Pichia pastoris can effectively metabolize galactose in Camellia oleifera oligosaccharides, significantly alleviating the symptoms of bee poisoning. This study provides technical support for improving honey bee metabolism using the bioengineering method, which is of great significance for promoting the sustainable development of the Camellia oleifera industry.
Source: Scientia Sinica Vitae
Authors: TianYu XU, Zhen LI, WeiYu YAN, XiaoBo WU, ZhiJiang ZENG
Language: English
Subfield: Insect Science
Domain: Life Sciences
SDG: Zero hunger
Open Access status: bronze
OpenAlex ID: W4414220448