Streptomyces griseus trypsin (EC 3.4.21.4) is one of the major extracellular proteinases, which is secreted by S. griseus. The coding gene of S. griseus trypsin (sprT) of S. griseus ATCC 10137 was cloned into the pUC 18. Transformants containing sprT were isolated from genomic library by using a synthetic oligonucleotide probe and characterized by direct gel hybridization and demonstrated a proteolytic activity in S. lividans 66. The intact proteinase gene could be delimited to a 1.8 kbp Bgl I fragment. The sodium dodecyl sulfate polyacrylamide gel electrophoreses of extracellular proteins produced by S. lividans 66 harboring pSGT, which is a pUJ718-2 derivative containing a 1.8 kbp Bgl I fragment, showed that the protein band responsible for the proteinase activity was located at the same position of the S. griseus trypsin purified from a pronase. Deduced amino acid sequence from the nucleotide sequence suggests that S. griseus trypsin is produced as a precursor, consising of three portions; an amino-terminal pre sequence (32 amino acid residues), a pro sequence (4 residues), and mature trypsin. The S. griseus trypsin cosists of 223 amino acids with a computed molecular weight of 23,112. The G+C content of the sprT if 72\%. The promoter-like sequence and a putative ribosomal binding site are followed by the coding sequence of S. griseus trypsin. Following a stop codon (TGA), a structure reminiscent of the Escherichia coli rho-independent transcriptional terminator exests. The predicted amino acid sequence of S. griseus trypsin differs from the published amino acid sequence [Olafson et al.(1975) Biochemistry 14, 1168-1177]. It should be corrected by the insertion of two serine residues near position 76(Ser) according to the numbering of $\alpha$-chymotypsin. The existence of the proline residues at pro and mature junction suggests that the processing of S. griseus trypsin is non-autocatalytic. This is the first report of the nucleotide sequence for a microbial ...