Sliding clamp-based affinity purification systems, methods of making and use thereof

Abstract

Described are affinity purification system that includes a carrier/surface that is non-cellular, and sliding clamp (SC) protein, and methods for purifying proteins that bind to the SC. The SC is associated with the carrier/surface via covalent/non-covalent interactions. To attain control of coupling site, the SC can be mutated via site-directed mutagenesis to introduce an exogenous residue and, the exogenous internal residue is conjugated to the non-cellular surface through the linker. The SC can also be coupled to the carrier via non-covalent interactions such as the affinity interactions involved in ligand/binding partner complex formation. The SC-based affinity purification system are used in a purification column as bait proteins, to isolate SC binding partners or non SC-binding proteins engineered to contain a SC binding site prior to its purification.

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