Purification and some properties of two boticins.

Abstract

Two bacteriocins (boticins) were elaborated without induction by strain S5, a nontoxigenic variant of Clostridium botulinum type E. After separation of the two active entities by gel filtration on Sephadex G-50, a large particle with boticin activity was isolated by density gradient ultracentrifugation, and a small soluble boticin was purified by continuous curtain electrophoresis and chromatography on sulfoethyl-Sephadex. Large and small boticins were purified 200- and 3,000-fold, respectively, with yields of 50% or more. The small boticin, a basic substance with molecular weight under 30,000, was the predominant species; the large boticin, a negatively charged particle with particle weight greater than 40 x 10(6), represented less than 20% of the total activity. Both purified boticins were resistant to heat and were attacked by proteolytic enzymes, but the large boticin was less thermostable and less sensitive to proteolytic enzymes than was the smaller variety. The activity of the large boticin was not reduced by treatment with urea or deoxyribonuclease. Both boticins exhibited sporostatic and bactericidal activities for C. botulinum type E, strain 070. A suspension of type E strain 070 vegetative cells was rendered nonviable within 9 min by the small boticin. The lethal action of this bacteriocin was not reversed by trypsin.