Efficient secretion of biologically active mouse tumor necrosis factor alpha by Streptomyces lividans

Abstract We have studied the production of mouse tumor necrosis factor α (mTNF) with Streptomyces lividans as host. m TNF cDNA was fused to the α-amylase-encoding gene ( aml ) of Streptomyces venezuelae ATCC15068 at 12 amino acids (aa) downstream from the signal-peptidase cleavage site so that the aa surrounding this processing site were conserved. S. lividans containing this construct secreted mTNF at moderately high levels (1–10 μg/ml) as a biologically active compound of high specific activity (1 × 10 8 units/mg protein). No unprocessed pre-protein and virtually no processed protein could be detected in the cell lysates. N-terminal aa sequence analysis indicated microheterogeneity (−3 to −6 forms) at the N-terminal site of secreted mTNF. It was demonstrated that this microheterogeneity was due to aminopeptidase activity.