We have compared the B-subunit pentamers of Escherichia coli heat-labile enterotoxin (LT) and verotoxin-1 (VT-1). The B-subunits of these bacterial toxins of the AB5 class have virtually no sequence identity and differ considerably in size (69 amino acids in VT-1 versus 103 in LT). They share a number of functional properties: pentamer formation, association with an A-subunit, binding to carbohydrate-containing lipids, and interaction with membranes. The structures of these proteins are very similar in some respects and very different in others. They can be superimposed with an rms deviation of only 1.29 A on the main chain atoms of 52 amino acids (0.98 A on 47 C alpha). Seven out of eight secondary structure elements are retained in the two toxins; only the N-terminal helix of LT is absent in VT-1. A disulfide bridge, which is essential for pentamer formation, is found in both structures, but in slightly different locations. However, the VT-1 B-subunit is much shorter on one side of the toxin, where the proposed membrane binding site of both VT-1 and LT is located. The monomer-monomer interface in the pentamer is much larger in LT than in VT-1, making the LT pentamer more stable. The central pores have a different character, and the sugar binding sites are not conserved between the toxins. The evolutionary relationship of the toxins is discussed.