Recombinant and truncated tetanus neurotoxin light chain: Cloning, expression, purification, and proteolytic activity

Fiorella Tonello, Rossella Pellizzari, Sebastiano Pasqualato, Guido Grandi, Evaristo Peggion, Cesare Montecucco

Research output: Contribution to journalArticle


Tetanus neurotoxin (TENT) consists of two disulfide-linked polypeptide chains, heavy (H) and light (L). The L chain is a zinc endopeptidase protein highly specific for vesicle-associated membrane protein (VAMP), which is an essential component of the exocytosis apparatus. Here we describe the cloning of the L chain of TeNT from Clostridium tetani strain Y-IV-3 (WS 15) and its expression in Escherichia coli as a glutathione S-transferase fusion protein. The full-length recombinant L chain, corresponding to residues 1-457, was obtained as a mixture of proteins of slightly different mass with identical N-terminal ends. To obtain a product useful for structural analysis and crystallization, a COOH-terminally truncated L chain (residues 1-427) was cloned, expressed, and purified with high yield. This truncated L chain is more active than the full-length and wild-type proteins in the hydrolysis of VAMP. Preliminary experiments of crystallization of the truncated recombinant L chain gave encouraging results.

Original languageEnglish
Pages (from-to)221-227
Number of pages7
JournalProtein Expression and Purification
Issue number2
Publication statusPublished - Mar 1999



  • Crystals
  • Metalloprotease
  • Tetanus
  • Tetanus neurotoxin

ASJC Scopus subject areas

  • Biochemistry

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