P-glycoprotein (P-gp) is an ATP-driven transmembrane transporter capable of effluxing a wide variety of structurally diverse and functionally unrelated hydrophobic compounds out of the cell. Multidrug resistance (MDR), often associated with the over-expression of P-gp, has been implicated as a major obstacle to effective chemotherapy for cancer, parasitic diseases, AIDS, and other diseases. Drug efflux mediated by P-gp is also involved in decreasing the oral bioavailability of drugs by limiting intestinal absorption. Our appreciation of the structural and functional aspects of P-gp has definitely improved in recent years, benefiting from the deciphering of the structure of some bacterial transporters that paved the way for construction of homology models for more complex transporters. Here, we will review the recent advances in the studies of the structure and functional characteristics of P-gp with the hopes of facilitating rational drug design in developing novel potent MDR modulators.