A Simple Method for Obtaining a Chaperone With a Variable Number of Non-Native Proteins Binding Sites

Proteins are one of the key elements of the cell. Their correct folding and prevention of aggregation under stress conditions are ensured by chaperone proteins. Understanding the principles of functioning of chaperones makes it possible to create and modify them to solve research problems or use them in biotechnology. We created an artificial chaperone based on the apical domain of GroEL. In this work, we present an approach that allows you to vary the number of non-native proteins binding sites on this chaperone. This approach allowed us to estimate the stoichiometry and binding strength of the non-native protein by the chaperone we created. It was shown that reliable binding of non-native αLA requires its interaction with several apical domains of GroEL. At the same time, the dissociation constant of such a complex does not change significantly with an increase in the number of binding domains in the oligomer. Up to 4 αLA molecules can be attached to a complete heptameric ring of apical domains. In the future, the proposed tool can also be used not only to study chaperones, but also to obtain proteins with a combination of any functional domains.