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Justin Siegel

Protein purification with Cobalt

Dick Selwood  Follow

Your question is about immobilized metal affinity chromatography (IMAC), often used to separate a protein expressed in the E. coli bacterium from other proteins made by it. The protein of interest is expressed with a histidine-containing affinity tag which binds to immobilized nickel or cobalt at neutral or basic pH. Immobilization is achieved by complexing to NTA (nitrilotriacetic acid) or IDA (iminodiacetic acid). For a discussion of the two ligands, see e.g. here.

There are a few proteins from E. coli (listed in this paper) that are known to bind along with the protein of interest. According to a post written by Damien Soghoian in 2004, nickel-NTA tends to bind stronger to histidine-tags while cobalt-NTA tends to bind more weakly, but this does not affect specificity (e.g. it would be across the board, not making the separation easier or harder).

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