Minicells, as compared with a whole cell preparation of a minA/B mutant of Escherichia coli, showed a number of changes in the structure of the murein sacculus. Minicell murein was enriched in LD-A2pm-A2pm crossbridges by about 66% and reduced in the amount of L-Ala-D-Glu dipeptide moieties by about 55%. In addition, the length distribution of the glycan strands in the murein was shifted to shorter lengths. In particular, the relative amount of the shortest possible strand, the size of a disaccharide, was more than doubled. Minicells were also found to have an altered penicillin-binding protein (PBP) pattern. Whereas PBP4 and PBP6 were greatly diminished, PBP8 was significantly increased. We consider it unlikely that the sort of changes observed in murein structure reflect the fact that minicells are composed of two hemispherical polar caps.