The Gram-positive bacterium, Bacillus subtilis and related species are widely used industrially as hosts for producing enzymes. These species possess a high potential to produce secreted proteins into the culture medium. Nevertheless, the secretion of heterologous proteins by these species is frequently inefficient. In this study, the human interferon-alpha 2b (hIFN-alpha 2b) was used as a heterologous model protein, to investigate the effect of B. subtilis AmyE propeptide in enhancing the secretion of heterologous proteins in B. subtilis. We found that the secretion production and activity of hIFN-alpha 2b with AmyE propeptide increased by more than threefold, compared to that without AmyE propeptide. The maximum amount of secreted hIFN-alpha 2b with propeptide was 14.8 +/- 0.6 mu g ml(-1). In addition, the pro-hIFN-alpha 2b bioactivity reached 5.4 +/- 0.5 x 10(7) U mg(-1), which is roughly the same level as that of the non-propeptide hIFN-alpha 2b. These results indicated that AmyE propeptide enhanced the secretion of the hIFN-alpha 2b protein from B. subtilis. This study provides a useful method to enhance the extracellular production of heterologous proteins in B. subtilis.