Modification and Production of Encapsulin

Encapsulins are a class of protein nanocages that are found in bacteria, which are easy to produce and engineer in E. coli expression systems. The encapsulin from Thermotoga maritima (Tm) is well studied, its structure is available, and without modification it is barely taken up by cells, making it promising candidates for targeted drug delivery. In recent years, encapsulins are engineered and studied for potential use as drug delivery carriers, imaging agents, and as nanoreactors. Consequently, it is important to be able to modify the surface of these encapsulins, for example, by inserting a peptide sequence for targeting or other functions. Ideally, this is combined with high production yields and straightforward purification methods. In this chapter, we describe a method to genetically modify the surface of Tm and Brevibacterium linens (Bl) encapsulins, as model systems, to purify them and characterize the obtain nanocages.