Acquired resistance to the aminoglycoside family of antibiotics has rendered this large and important family of compounds virtually unusable. Resistance is primarily mediated by three classes of enzymes, typically residing on transposable elements in resistant bacteria. These enzymes, the phosphotransferases, acetyltransferases and adenyltransferases, chemically modify the aminoglycosides, which either interferes with drug transport or the binding of the drug at the site of antibacterial action, the 30S ribosomal subunit. The structures of several members of the aminoglycoside-modifying enzyme family are now known, and it is hoped that through a better understanding of these enzymes, both from a structural and mechanistic view-point, could lead to the development of either rationally-designed novel aminoglycosides, or specific structure-based enzyme inhibitors. Such developments could help to bring these compounds back to the forefront of modern antimicrobial chemotherapy. This review focuses on the structural details of the enzymes whose crystal structures are known and on the implications of these findings for devising novel strategies to overcome resistance to this broad class of antibiotics.