Colicin E4 has been iodinated with ['251]iodine. The relationship between the degree of iodination and the biological activity has been determined. In contrast to colicins E2, E3, Ia, Ib or cloacin, conventional saturation curves with sensitive cells are not obtained with colicin E4. A turnover ofcolicin on the receptors is evidenced. Analysis of the colicin molecules (M, 56000 2000) incubated in buffer with sensitive cells, on sodium dodecylsulfate/polyacrylamide gels shows that cleavage of the colicin molecules occurs to yield two fragments of 27000 M , and 26000 M, respectively. Strains selected for resistance to colicin E4 are defective in their ability to cleave this bacteriocin, whereas tolerant strains, which have an intact receptor but are not killed by colicin E4. cleave like sensitive cells. Colicin -receptors interaction, which results in cleavage, is much less sensitive to the degree of iodination than the biological activity. Kinetics of cleavage as well as the effect of various treatments of sensitive cells are described. Among the various parameters which alter the cleavage, the most interesting is the ionic strength effect. In the presence of 0.5",, or 1 0 o NaC1. colicin E4 cleavage practically is not evidenced. We suggest that the equilibrium constant of the colicin-receptor complex is very sensitive to ionic strength. The mechanism of colicin E4 cleavage with regard to the killing action is discussed.The early hypothesis presented by Fredericq in 1946 [ l ] which postulated that bacteria sensitive to a given colicin contain a specific receptor substance on their surface, which serves as the site of attachment for the bacteriocin, has been fully confirmed in recent years. There is a wide agreement that a first step in the killing of Escherichirr coli by colicins is the adsorption of colicin molecules to receptor sites on the bacterial surface. In the case of colicins E2, €3 [2], Ia and Ib [3,4] and of cloacin [5], binding of radioactive colicins to cells was measured directly. In the accompanying study [6] we described the purification and the molecular properties of colicin E4. This bacteriocin causes physiological effects similar to those caused by colicins E l , K , Ia, Ib and L-JF246. The aim of this work was to study the conditions and the effects of the interaction of colicin E4 with its receptor site. We present evidence which suggests that, in contrast to all colicins previously studied [7], colicin E4 undergoes a turnover on the receptor sites triggering a proteolytic activity cleaving the colicin molecules like a substrate and releasing two fragments devoid of h-rm,. Lactoperoxidase (EC 1 .1 1.1 .7).
