Excretion of an extracellular protease of Serratia marcescens ATCC 25419 occurred during logarithmic growth and was highest (per cell) when cultures reached the stationary growth phase. Production of the extracellular protease was induced by leucine or casein in minimal medium or by growth in tryptone-yeast medium. In the late stationary phase an intracellular protease activity accumulated which was also observed in mutants with very low extracellular protease activity. The excreted protease was the dominant protein in the growth medium. The protease was purified to homogeneity by column chromatography on Bio-Gel P-100 and on DEAE-cellulose. Quantitative amino acid analysis revealed the absence of sulfur-containing amino acids. The enzyme consists of one polypeptide chain. A molecular weiht of 51,000 and 55,000 was estimated using polyacrylamide gel electrophoresis and chromatography on Bio-Gel P-100 respectively. The enzyme cleaved only N-alpha-benzoyl-DL-lysine-and-arginine-nitroanilides but not the corresponding leucine or tyrosine derivatives nor a set of di- and tripeptides.