α-葡萄糖苷酶抑制剂高通量筛选模型的建立及其应用

Establishment and application of a high-throughput model for screening α-glucosidase inhibitors

摘要:

摘要：【目的】针对人α-麦芽糖苷酶这个糖代谢途径中重要的靶蛋白，建立α-糖苷酶抑制剂高通量筛选模型。【方法】采用毕赤酵母表达系统克隆和表达人α-麦芽糖苷酶.利用酶的催化特性建立α-糖苷酶抑制剂筛选模型.应用该模型对放线菌代谢产物库进行高通量筛选.通过构建16S rRNA系统发育树分析阳性菌株的分类地位。【结果】首次成功克隆、表达了具催化活性的人α-麦芽糖苷酶N端结构域.针对人α-麦芽糖苷酶N端催化结构域，建立α-糖苷酶抑制剂的筛选模型.对包含近2000株放线菌代谢产物的天然产物库进行高通量筛选，最终得到20株α-麦芽糖苷酶抑制剂生产菌株.其中19株放线菌为链霉菌属，且在分类学上具有丰富的多样性。【结论】本研究建立的α-糖苷酶抑制剂高通量筛选模型具有很强的实用价值，可用于新型糖苷酶抑制剂类降糖药物的开发。

Abstract:

Abstract: [Objective] Targeted at the important enzyme in human glucose metabolic pathway, the purpose of this paper is to establish α-glucosidase inhibitors high throughput screening model. [Methods] Pichia pastoris expression system was used to clone and express the human α-maltase glucosidase. Using the catalytic properties of enzyme to establish α-glucosidase inhibitor screening model. This model was applied in screening of actinomycete metabolites library. The taxonomic status of positive strains were analyzed by constructing 16S rRNA phylogenetic tree. [Results] The N-terminal catalytic domain of human α-maltase glucosidase was successfully cloned and expressed for the first time. The high-throughput screening model of α-glucosidase inhibitors was established. A natural product library containing metabolites from nearly 2000 actinomycetes was screened, 20 α-maltase glucosidase inhibitor producing strains were obtained finally, of which, 19 strains initially identified as Streptomyces, and showed taxonomically rich diversity. [Conclusion] The α-glucosidase inhibitor high-throughput screening model has high practical value, this work laid the foundation for developing new hypoglycemic drugs.