Expression of catalytic antibodies in eukaryotic systems

Захаров, А. В.; Смирнов, Иван; Serebryakova, Marina V.; Dronina, M. A.; Kaznacheeva, A. V.; Kurkova, I. N.; Belogurov, Alexey A.; Friboulet, Alain; Пономаренко, Н. А.; Gabibov, Alexander; Бобик, Т. В.

doi:10.1134/s0026893311010171

Cited by 8 publications

(3 citation statements)

References 22 publications

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“…Recombinant FabA17 containing or light-chain constant regions was produced in the methylotrophic yeast Pichia pastoris GS115 using the modified expression vector pPICZA/Jk1 (Zakharov et al, 2011) based on pPICZA (Invitrogen, USA) ( Supplementary Fig. S1 1 ).…”

Section: Protein Expression and Purificationmentioning

confidence: 99%

“…Expression constructs for HC H (human Ab heavy chain containing the C H1 constant domain with hinge region) and A17 light chain (Zakharov et al, 2011) and the constant region of human 2 light chain with the J2 (joining) segment (Gabibov et al, 2011) were prepared as described in the respective studies. To construct pPICZA/J2, the PCRamplified DNA fragment corresponding to J22 was digested with KpnI and SacII and ligated into pPICZA/Jk1 at the appropriate restriction sites.…”

Section: Protein Expression and Purificationmentioning

confidence: 99%

“…To express full-length A17 reactibody in CHO cells, we used vectors permitting the production of a corresponding Ab with human constant domains of subclass IgG1/ (Smirnov et al, 2011). Some differences were observed in the catalytic efficiency of full-length A17 or its Fab fragment compared with the parent scFv molecule (Zakharov et al, 2011). On one hand, these differences could result from structural stabilization of the active centre owing to the presence of additional constant domains.…”

Section: Molecular Organization Of the A17 Reactibodymentioning

confidence: 99%

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Role of κ→λ light-chain constant-domain switch in the structure and functionality of A17 reactibody

Ponomarenko

Chatziefthimiou

Kurkova

et al. 2014

Acta Cryst D Biol Crystallogr

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Section: Protein Expression and Purificationmentioning

confidence: 99%

Section: Protein Expression and Purificationmentioning

confidence: 99%

Section: Molecular Organization Of the A17 Reactibodymentioning

confidence: 99%

See 1 more Smart Citation

Role of κ→λ light-chain constant-domain switch in the structure and functionality of A17 reactibody

Ponomarenko

Chatziefthimiou

Kurkova

et al. 2014

Acta Cryst D Biol Crystallogr

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New Genetic Constructs for Generation of Stable Therapeutic Antibodies to Organophosphorus Toxins in Methylotrophic Yeasts Pichia Pastoris

et al. 2016

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We propose a new method of obtaining of stable Fab-fragments of antibodies in Pichia pastoris expression system. Recently, we obtained Fab-fragments of antibodies neutralizing organophosphorus toxins. However, high yield of the target products was not attained because of high level of proteolytic degradation. In the present study, we identified sites of proteolytic degradation in Fab-fragments and endogenous proteases performing degradation, which allowed obtaining optimized genetic constructs for expression of antibody heavy chains (IgGγ1) and kappa and lambda isotypes of light chains. Co-transformation of these vectors allowed obtaining Fab-fragments of antibodies to organophosphorus toxins without proteolytic degradation of the product.

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Genetic Engineering of Native Chain Combinations of B-Cell Repertoires on the Surface of Methylotrophic Yeasts Pichia pastoris

et al. 2017

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We designed genetic constructs for exposing Fab-fragment library of natively paired single cell B-cell receptors on the surface of Pichia pastoris yeast cells. We have previously obtained the A17 antibody in our laboratory [6]. In this study we showed that the newly designed genetic constructs provide a compatible level of A17 antibody Fab fragment on the surface of yeast cells as well as in the case of vectors containing DNA fragments corresponding to each chain of the antibody. The data suggest that the developed approach for constructing immunoglobulin gene libraries is adequate and fully convenient for studying properties of the real human B-lymphocyte repertoire.

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Expression of catalytic antibodies in eukaryotic systems

Cited by 8 publications

References 22 publications

Role of κ→λ light-chain constant-domain switch in the structure and functionality of A17 reactibody

Role of κ→λ light-chain constant-domain switch in the structure and functionality of A17 reactibody

New Genetic Constructs for Generation of Stable Therapeutic Antibodies to Organophosphorus Toxins in Methylotrophic Yeasts Pichia Pastoris

Genetic Engineering of Native Chain Combinations of B-Cell Repertoires on the Surface of Methylotrophic Yeasts Pichia pastoris

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