Dissociation and Reconstitution of Colicin E3 and Immunity Substance Complex

Abstract

It has recently been found that so-called native colicin E3, which has been used for studies of its mode of action, is a complex of two kinds of proteins. The complex could be dissociated into the two components in SDS. These components were isolated by gel filtration in 1% SDS followed by treatment with Dowex-2 to remove bound SDS. One component, characterized by its low molecular weight, prevented colicin E3-induced inhibition of poly(U)-dependent protein synthesis and was designated as immunity substance. The other component (protein A), which was of high molecular weight, had 100-fold higher in vitro ribosome-inactivating activity than native colicin E3, but had lower bacteriocidal activity.
Colicin E3 was reconstituted from the two isolated protein components. The reconstituted colicin E3, when compared with protein A, showed a decrease in in vitro activity (inhibition of poly(U)-dependent protein synthesis), but had higher bacteriocidal activity in vivo. Thus complex formation of protein A with immunity substance should play an important role in the bacteriocidal action, but protein A itself might inactivate ribosomes in the interior of the sensitive cells.