Many heterologous polypeptides fail to fold into their native state when expressed in Escherichia coli; instead, they are either degraded by the cellular proteolytic machinery or accumulate in insoluble form, typically as inclusion bodies. Misfolding is a particularly vexing problem in the expression of mammalian proteins, especially those that are composed of multiple subunits, have several disulfide bonds, or contain prosthetic groups. Fortunately, bacteria exhibit a remarkable physiological plasticity that can be successfully exploited to improve protein folding. Significant yields of active heterologous proteins have been obtained through strategies that include the co-expression of homologous or heterologous folding accessory proteins, the optimization of growth conditions, and the use of fusion proteins. A flood of recent reports documenting the successful production of complex eukaryotic proteins in active form have demonstrated that bacteria can provide the proper environment for the folding of the vast majority of recombinant polypeptides.