title: Proteolysis of human tumor necrosis factor (TNF) by endo- and exopeptidases: process of proteolysis and formation of active fragments.
abstract: Recombinant human tumor necrosis factor (TNF) was digested with endopeptidases under mild conditions. Incubation of the TNF (155-amino-acid TNF) with trypsin, Staphylococcus aureus V-8 protease or chymotrypsin initially released small peptides derived from the amino (N)-terminal region of TNF, but did not release peptides from the carboxyl (C)-terminal region. The TNF was resistant to carboxypeptidases A and Y under a non-denaturing condition, but in the presence of urea or sodium dodecyl sulfate the C-terminal amino acid was released quantitatively by these peptidases. These results indicate that the N-terminal region of the TNF molecule is accessible to protease, while the C-terminal region is not susceptible to degradation. When the TNF was incubated with seven kinds of endopeptidases, its activity rapidly disappeared. At an early stage of the degradation, one active fragment was detected among the fragments produced with trypsin or pronase P, but no active fragments were detected on the degradation with the other peptidases. The active fragment was a fragment lacking the four N-terminal amino acid residues of the TNF. These results suggest that TNF is initially degraded at the N-terminal region by an endopeptidase and loses its activity as the degradation proceeds.
authors: Department of Pharmacology I, Dainippon Pharmaceutical Co., Ltd., Osaka, Japan.
authors: K
authors: K
authors: Nakamura
authors: M
authors: M
authors: Komiya
mesh_terms: D000595:Amino Acid Sequence; D002268:Carboxypeptidases; D010450:Endopeptidases; D004926:Escherichia coli; D006801:Humans; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000302:isolation & purification; D011402:Pronase; D011994:Recombinant Proteins / Q000737:chemistry; D014357:Trypsin; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry*
chemical_list: D010446:Peptide Fragments; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D002268:Carboxypeptidases; D010450:Endopeptidases; D014357:Trypsin; D011402:Pronase