Synthesis and secretion of hirudin by Streptomyces lividans.
To examine the secretory production of heterologous proteins by Streptomyces lividans, we fused the DNA encoding the signal peptide of the alpha-amylase inhibitor tendamistat, derived from S. tendae with a synthetic gene encoding the thrombin inhibitor hirudin. The analysis of secretion by immunoblots revealed an efficient translocation of hirudin through the membrane, with no detectable immunoreaction among the cellular proteins. The secreted hirudin was stable in the shaking culture for about 6 days. A comparison of the hirudin secreted by S. lividans and recombinant reference hirudin from yeast by immunoblots and thrombin inhibition assays shows that hirudin from Streptomyces has a lower specific activity, which may be due to a different aminoterminal sequence or to inexact processing of the precursor.
Bender
E
E
Institut für Organische Chemie, Universität Frankfurt, Main, Federal Republic of Germany.
Vogel
R
R
Koller
K P
KP
Engels
J
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D006629:Hirudins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics; D010957:Plasmids; D021382:Protein Sorting Signals / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013302:Streptomyces / Q000235:genetics*; D013917:Thrombin / Q000037:antagonists & inhibitors; D000516:alpha-Amylases / Q000037:antagonists & inhibitors
D006629:Hirudins; D010455:Peptides; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D000516:alpha-Amylases; D013917:Thrombin; C030526:tendamistate