Streptomyces lividans is known to produce large amounts of proteins in culture supernatants. In this report, to expand the secretory expression system with a strong promoter derived from phospholipase D of Streptoverticillium cinnamoneum, we expressed three kinds of proteins: transglutaminase from Stv. cinnamoneum (StvcMTG) and β-1,4-endoglucanase and β-glucosidase from Thermobifida fusca YX . The StvcMTG gene was introduced into S. lividans using the shuttle vector pUC702 for Escherichia coli and S. lividans, and high level secretory production of StvcMTG (230 μg/ml in the culture supernatant) was achieved. The other prokaryotic proteins, β-1,4-endoglucanase and β-glucosidase, were also expressed in (His) 6-tag fused form into culture supernatants and retained high activity. Furthermore, complete purification was achieved by conventional column or affinity column chromatography for each recombinant protein with 1 mg/ml over protein concentration. Three independent proteins were thus successfully expressed and purified, and we expect to use this system for the expression of other valuable heterologous proteins.