Title: Expression, purification and comparative characterisation of enzymatically deactivated recombinant botulinum neurotoxin type A

Authors: Weiping Yang, Paul Lindo, Stephen Riding, Tzuu-Wang Chang, Shuowei Cai, Thuan Van, Roshan Kukreja, Yu Zhou, Kruti Vasa, Bal Ram Singh

Addresses: Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA. ' Botulinum Research Center, Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, North Dartmouth, MA 02747-2300, USA

Abstract: Botulinum Neurotoxin (BoNT) is responsible for botulism, a severe and often deadly disease. Light chain of BoNT behaves as endopeptidase, cleaving the SNARE proteins, and inhibits the neurotransmitter release. A double-mutant E224A/E262A full-length BoNT Type A was successfully cloned and expressed in E. coli. The purified protein lacks the endopeptidase activity involved in the toxic action, and is structurally compatible with the native BoNT/A. Thus this molecule not only can be used as a safe surrogate for study of BoNT, but also can be potentially used as an antidote against botulism, and as a vaccine candidate for botulism.

Keywords: botulinum neurotoxins; clostridium; endopeptidase; mutant BoNT; structure; toxicity; botulism; antidote; botulism vaccines; E. coli; BoNT surrogate; cloning.

DOI: 10.1504/TBJ.2008.026477

The Botulinum Journal, 2008 Vol.1 No.2, pp.219 - 241

Published online: 16 Jun 2009 *

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