Bacitracin: substantiation and elimination of contaminating proteolytic activity and use as an affinity chromatography ligand to purify a siderophore-degrading enzyme.

Abstract

Bacitracin is a commercial general peptidase inhibitor that may be used to purify proteases. Significant protease contamination of a commercial bacitracin preparation was noted and four procedures were developed to overcome the contamination. Dialysis, gel-filtration chromatography, molecular weight cutoff filters, and heat inactivation were effective, resulting in the diminution or elimination of proteolysis while maintaining the inhibitory effect of bacitracin. Attachment of bacitracin to an affinity chromatography resin did not immobilize a siderophore-degrading enzyme, as has been noted with peptidases. It did, however, result in its partial purification from some of the contaminating proteins originally present.