6-Deoxyerythronolide B, the macrocyclic aglycone of the antibiotic erythromycin, is synthesized by a polyketide synthase (PKS) that has emerged as the prototypical modular megasynthase. A variety of molecular biological, protein chemical, and biosynthetic experiments over the past two decades have yielded insights into its mechanistic features. More recently, high-resolution structural images of portions of the 6-deoxyerythronolide B synthase have provided a platform for interpreting this wealth of biochemical data, while at the same time presenting a fundamentally new basis for the design of more detailed investigations into this remarkable enzyme. For example, the critical roles of domain-domain interactions and nonconserved linkers, as well as large interdomain movements in the structure and function of modular PKSs, have been highlighted. In turn, these insights point the way forward for more sophisticated and efficient biosynthetic engineering of complex polyketide natural products.