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Camelid‐derived heavy‐chain nanobody against Clostridium botulinum neurotoxin E in Pichia pastoris
Author(s) -
Baghban Roghayyeh,
Gargari Seyed Latif Mousavi,
Rajabibazl Masoumeh,
Nazarian Shahram,
Bakherad Hamid
Publication year - 2016
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1002/bab.1226
Subject(s) - pichia pastoris , botulism , antitoxin , escherichia coli , clostridium botulinum , biology , recombinant dna , yeast , antibody , pichia , epitope , microbiology and biotechnology , immunoglobulin light chain , toxin , biochemistry , gene , genetics
Botulinum neurotoxins (BoNTs) result in severe and often fatal disease, botulism. Common remedial measures such as equine antitoxin and human botulism immunoglobulin in turn are problematic and time‐consuming. Therefore, diagnosis and therapy of BoNTs are vital. The variable domain of heavy‐chain antibodies (VHH) has unique features, such as the ability to identify and bind specifically to target epitopes and ease of production in bacteria and yeast. The Pichia pastoris is suitable for expression of recombinant antibody fragments. Disulfide bond formation and correct folds of protein with a high yield are some of the advantages of this eukaryotic host. In this study, we have expressed and purified the camelid VHH against BoNT/E in P. pastoris . The final yield of P. pastoris ‐expressed antibody was estimated to be 16 mg/l, which is higher than that expressed by Escherichia coli . The nanobody expressed in P. pastoris neutralized 4LD 50 of the BoNT/E upon i.p. injection in 25% of mice. The nanobody expressed in E. coli extended the mice's survival to 1.5‐fold compared to the control. This experiment indicated that the quality of expressed protein in the yeast is superior to that of the bacterial expression. Favorable protein folding by P . pastoris seems to play a role in its better toxin‐binding property.

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