The catalytic and resistance mechanisms of bacterial dihydropteroate synthase (DHPS), the target of the sulfa drug antibiotics, have finally been solved thanks to work by Yun et al. Sulfonamides were among the first antibiotics discovered and are potent against bacteria and some eukaryotic pathogens, but resistance arising through mutations in DHPS has severely limited use of these drugs in recent years. DHPS is a key enzyme in the folic acid biosynthesis pathway, and it catalyzes the production of the folate intermediate 7,8-dihydropteroate from 6-hydroxymethyl-7,8-dihydropterin pyrophosphate and p-aminobenzoic acid (PABA). Sulfa drugs act by mimicking PABA and binding to DHPS. First, the authors used computational predictions of enzyme reactions and crystallization of DHPS enzymes from Bacillus anthracis and Yersinia pestis to establish an essential role for bound magnesium in catalysis. Then, using structural enzymology techniques that take advantage of the fact that many proteins retain catalytic activity in a crystalline form, and by soaking DHPS crystals from Y. pestis in substrates and substrate analogs, the authors deduced the complete catalytic mechanism, including where DHPS contacts sulfa drugs. This should pave the way for rational design of new sulfa drugs. (Science 335, 1110–1114, 2012)