Human drug transporters often play key roles in determining drug accumulation within cells. Their activities are often directly related to therapeutic efficacy, drug toxicity as well as drug–drug interactions. However, the progress for interpretation of their crystal structures is relatively slow. Hence, conventional biochemical studies together with computer modeling became useful manners to reveal essential structures of these membrane proteins. Over the years, quite a few structure–function relationship information had been obtained for members of the two major transporter families: the ATP-binding cassette family and the solute carrier family. Critical structural features of drug transporters include transmembrane domains, post-translational modification sites and domains for cell surface assembly and protein–protein interactions. Alterations at these important sites may affect protein stability, trafficking to the plasma membrane and/or ability of transporters to interact with substrates.