Binding domains of colicins E1, E2 and E3 in the receptor protein BtuB of Escherichia coli

Eight reagents specifically modifying amino acids were applied to cells of a standard Escherichia coli colicin indicator strain to follow in vivo chan ges of its binding capacity for colicins E1-E3 and hence the binding domain s (epitopes) for them in the outer membrane receptor protein BtuB. The effe ct of these reagents was also investigated in a mutant strain carrying an e xtensive BtuB deletion. The following differences of the binding epitopes c ould be ascertained. Colicin E1. Blockage of OH-groups, just as N-substitut ion of His and modification of Arg and Trp enhance binding of colicin E1. I n the deleted receptor, also abolition of carboxylic anion bonds enhances i ts affinity for colicin E1. it follows that colicin E1 is bound, most of al l, to the hydrophobic domain A (loops 1 + 2) of BtuB. Colicins E2 and E3 bo th exert rather analogous binding parameters. In contrast to E1, O-substitu tion of Ser and Thr dramatically decreases the E2 and E3 binding, similarly to modification of Lys. There is also a clear difference in the binding af finity of the domain for E2 and/or E3 and for E1 following modifications of their Arg and His. Colicins E2 and E3 are rather bound to the hydrophilic domain B (loops 5-7) of the receptor. In this respect, interactions of coli cins E2 and E3 with deeper parts of A and B domains (Trp, several Arg, Lys and His residues) exhibited subtle differences. Acidic pH (4.5-6.0) shows a positive, while pH 7.0-8.5 a rather negative impact on the receptor-bindin g function for the colicins. It was clearly demonstrated that there is just a partial difference between the binding behavior of colicins E1, E2 and/o r E3.