The effect of the hexahistidine-tag in the oligomerization of HSC70 constructs

The hexahistidine is a fusion tag used for the isolation of proteins via an immobilized metal-ion affinity chromatography (IMAC). In the present study, we have purified and analyzed two constructs of the heat shock protein HSC70 in the presence or the absence of the His-tag (C30WT-His+/C30WT and C30ΔL-His+/C30ΔL). The oligomerization properties of the constructs were analyzed by size exclusion chromatography (SEC) and analytical ultracentrifugation (AU). Results from SEC analysis indicated that the His-tag promotes the dimerization of C30ΔL-His+ but has no effect on the elution profile of C30WT-His+, compared to their respective untagged forms C30ΔL and C30WT. These observations were also confirmed by AU analysis which indicates that C30ΔL is stabilized in the dimeric form in the presence of the His-tag. These results emphasize the need to remove the His-tag before structural characterization of some recombinant proteins.