The targets of β-lactam antibiotics are bacterial DD-peptidases that catalyze the final steps of peptidoglycan biosynthesis. Bacterial resistance to β-lactams is achieved by the production of β-lactamases, enzymes that catalyze β-lactam hydrolysis. Structural studies of both of these groups of enzymes, their substrates and of β-lactams have led to the conclusion that β-lactamases have evolved from a DD-peptidase ancestor. Thus, the active sites of DD-peptidases and serine β-lactamases are very similar. Why is it then that the active site of a serine β-lactamase can catalyze hydrolysis of a β-lactam while that of a DD-peptidase cannot? In view of the active site similarities, why was it necessary for β-lactamases to evolve at all? The aim of this review is to examine our current understanding of these issues in terms of the crystal structures of the relevant enzymes that are now available, rounding off the analysis with speculation where necessary.