抗CEACAM5纳米抗体在大肠杆菌中的高效周质表达

Efficient Expression of Anti-CEACAM5 Nanobody in Escherichia coli Periplasm

纳米抗体具有相对分子质量小、稳定性高、特异性强以及能够穿过血脑屏障等众多优势,而且其能够通过大肠杆菌、酵母等简单微生物大量表达。大肠杆菌周质表达纳米抗体是目前纳米抗体制备主要的方法之一,但是纳米抗体的表达水平还相对较低。以pET22b和pMES4为载体构建了两种重组表达质粒,并且比较了他们在不同宿主菌中表达纳米抗体的情况。结果表明,以pMES4为载体的大肠杆菌能够在周质中可溶性表达纳米抗体,而以pET22b为载体的大肠杆菌表达的纳米抗体无法分泌到周质空间中。随后,通过IPTG浓度优化及5 L发酵罐过程,控制实现了纳米抗体在大肠杆菌周质中的表达量达到308. 32 mg/L。研究提供了一种高效表达纳米抗体的方法,为纳米抗体的规模化制备奠定了技术基础。

Nanobodies offer multiple advantages over conventional antibodies in terms of size,stability,solubility,immunogenicity,and production costs,with improved tumor uptake and blood clearance.Additionally,the recombinant expression of nanobodies is robust in various expression systems,such as Escherichia coli,Saccharomyces cerevisiae and Pichia pastoris.Periplasmic expression of nanobody was one of main nanobody production ways in E.coli.However,level of nanobody expression was low in E.coli periplasm.In the study,two recombinant expression vectors,pET22b-2D5 and pMES4-2D5,were constructed for nanobody expression in E.coli periplasm.The results indicated that nanobody 2D5 could be efficiently expression in E.coli periplasm with pMES4-2D5 vector.With optimizations,a high nanobody yield of 308.2 mg/L was achieved in a 5L-fermentor.The results of this study provide an efficient way to produce nanobody,and laid a technical foundation for scale-production of nanobodies.