Ultra-High-Throughput Directed Evolution of Antimicrobials using Droplet Microfluidics (Ref: 3976)

抗菌素耐药性已演变为一个主要的卫生保健威胁，而正在开发的抗生素数量的减少进一步加剧了这一威胁。目前正在积极研究噬菌体和噬菌体内溶素（它们的分解酶）等抗生素的替代品。该博士项目旨在了解内溶素活性的序列-功能关系，并揭示成功细菌溶解的规则。在这个项目中，我们将把微流控液滴的尖端高通量筛选技术与蛋白质工程相结合，创建一个内溶素进化平台。微滴技术在高通量、低成本的酶进化和序列空间探索方面已经显示出巨大的潜力。每个液滴，其体积通常在毫升到纳升之间，代表了单个突变酶被限制的单一反应。通过封装对应于单个酶变体的单个基因，可以以时间和资源效率的方式筛选大型基因库。总之，这是一个开发新型高通量功能分析的实验项目，可以使用微流体、光学询问装置、蛋白质工程、基因表达和微生物学的组合来识别新型的溶菌酶。

Antimicrobial resistance has evolved into a major healthcare threat which is further exacerbated by the diminished number of antibiotics in development. Alternatives to antibiotics such as phages and phage endolysins (their lytic enzymes) are being actively researched. This PhD project aims at understanding sequence-function relationships for endolysin activity and unravel the rules for successful bacteriolysis.In this project, we will combine cutting-edge high-throughput screening technologies in microfluidic droplets with protein engineering to create an endolysin evolution platform. Microdroplet technology has already shown enormous potential for enzyme evolution and the exploration of sequence space at high-throughput and low cost. Each droplet, whose volume typically ranges from femto to nanoliter scale represents a single reaction in which individual mutant enzymes are confined. By encapsulating a single gene corresponding to a single enzyme variant, large gene libraries can be screened in a time and resource efficient manner. In summary, this is an experimental project to develop novel high-throughput functional assays that can identify novel bacteriolytic enzymes using a combination of microfluidics, optical interrogation setups, protein engineering, gene expression and microbiology.