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Expression in Pichia pastoris of a recombinant scFv form of MAb 107, an anti human CD11b integrin antibody

Abstract : We herein report the production in the methylotrophic yeast Pichia pastoris of a recombinant scFv form of monoclonal antibody 107 [MAb 107]. This antibody is directed to the leukocyte adhesion molecule CR3 (CD11b/CD18) and is specific of the CD11bA domain. MAb 107 is a ligand mimic of CR3 that blocks the adhesion of leukocyte and their transendothelial migration (diapedesis), a feature that confers this MAb a potential anti inflammatory activity. Expression of scFv 107 as a myc-(His)6 secreted fusion protein was achieved in the yeast P. pastoris using plasmid pICZαB. A spontaneous specific cleavage of the myc-His tag was consistently observed regardless of the culture conditions and the addition of different concentrations of casaminoacids in the medium. Loss of the myc-His tag did not affect the binding activity of scFv 107. This loss is due either to a specific proteolysis or to a mechanical cleavage. Optimization of the production of scFv 107 in shake flasks was carried out using an L16 array Taguchi design. A level of purified scFv-myc-His fusion protein estimated at 40 mg/L was reached at pH 6 and a temperature of 27 °C. The production of large amounts of recombinant scFv form of MAb 107 is essential for further molecular studies of the interactions between CR3 and its ligands, mainly those mediated by the CD11b A domain. It will also facilitate the investigation of its anti inflammatory activity in vivo.
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Contributor : AbdelHakim Ben Hassine Connect in order to contact the contributor
Submitted on : Thursday, February 28, 2019 - 12:04:49 PM
Last modification on : Thursday, May 16, 2019 - 10:45:55 AM

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Naouel Guedel Ben Tanfous, Héla Kallel, M. Ali Jarboui, Dahmani M. Fathallah. Expression in Pichia pastoris of a recombinant scFv form of MAb 107, an anti human CD11b integrin antibody. Enzyme and Microbial Technology, Elsevier, 2006, 38 (5), pp.636-642. ⟨10.1016/j.enzmictec.2005.07.014⟩. ⟨pasteur-02052114⟩



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