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http://dx.doi.org/10.5352/JLS.2020.30.11.1012

The Improved Antigen-binding Activity of Biosimilar Remicade ScFv Antibodies by Fusion of the Leucine Zipper Domain  

Kim, Jin-Kyoo (Division of Biohealth Science, College of Natural Sciences, Changwon National University)
Kim, Tae Hwan (Division of Biohealth Science, College of Natural Sciences, Changwon National University)
Publication Information
Journal of Life Science / v.30, no.11, 2020 , pp. 1012-1020 More about this Journal
Abstract
Remicade is a therapeutic biosimilar natural antibody in which the mouse variable domain has been linked to the human constant domain. It is a chimeric monoclonal antibody specific to tumor necrosis factor-alpha (TNF-α) and has been developed for the treatment of rheumatoid arthritis. To investigate the biological activity of the Remicade antibody, we carried out a bioinformatics study using a protein data bank to characterize the TNF-α antigen binding mechanism of the Remicade natural antibody. Because the production of the Remicade antibody is often limited by genetic instability of the natural antibody-producing cell, we generated a Remicade single-chain variable domain fragment antibody (Remicade) in which a heavy chain variable domain (VH) is joined with a light chain variable domain (VL) by a polypeptide linker. Furthermore, Remicade was fused to a leucine zipper (RemicadeScZip) for higher production and higher antigen-binding activity than Remicade. The Remicade and Remicade ScZip were expressed in Escherichia coli and purified by a Ni+-NTA-agarose column. As expected, the purified proteins had migrated as 28.80 kDa and 33.96 kDa in sodium dodecyl sulfate-polyacrylamide electrophoresis. The TNF-α antigen binding activity of Remicade was not observed by ELISA and western blot. In contrast, RemicadeScZip showed antigen-binding activity. Additional bio-layer interferometry analysis confirmed the antigen-binding activity of RemicadeScZip, suggesting that the leucine zipper stabilized the folding of RemicadeScZip in a denatured condition and improved the TNF-α antigenbinding activity.
Keywords
Bio-layer interferometry; leucine zipper; remicade; remicadescZip; single-chain variable domain fragment;
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1 Bondeson, J. and Maini, R. 2001. Tumour necrosis factor as a therapeutic target in rheumatoid arthritis and other chronic inflammatory diseases: the clinical experience with infliximab (REMICADE). Int. J. Clin. Pract. 55, 211-216.
2 Bong, K. T. and Lee, H. 2020. Analysis and estimation for market share of biologics based on google trends big data. J. Soc. Korea Ind. Syst. Eng. 43, 14-24.   DOI
3 Carmona, L., Gomez-Reino, J. J., Rodriguez-Valverde, V., Montero, D., Pascual-Gomez, E., Mola, E. M., Carreno, L., Figueroa, M. and Group, B. 2005. Effectiveness of recommendations to prevent reactivation of latent tuberculosis infection in patients treated with tumor necrosis factor antagonists. Arthritis Rheum. 52, 1766-1772.   DOI
4 Chang, H. C., Bao, Z. Z., Yao, Y., Tse, A., Goyarts, E. C., Madsen, M., Kawasaki, E., Brauer, P. P., Sacchettini, J. C. and Nathenson, S. G. 1994. A general method for facilitating heterodimeric pairing between two proteins: application to expression of alpha and beta T-cell receptor extracellular segments. Proc. Natl. Acad. Sci. USA. 91, 11408-11412.   DOI
5 Chothia, C. and Lesk, A. M. 1987. Canonical structures for the hypervariable regions of immunoglobulins. J. Mol. Biol. 196, 901-917.   DOI
6 De Kruif, J. and Logtenberg, T. 1996. Leucine zipper dimerized bivalent and bispecific scFv antibodies from a semi-synthetic antibody phage display library. J. Biol. Chem. 271, 7630-7634.   DOI
7 Han, S. H. and Kim, J. K. 2016. The development of anti-DR4 single-chain Fv (ScFv) antibody fused to Escherichia coli alkaline phosphatase. Kor. J. Microbiol. 52, 10-17.   DOI
8 Huston, J. S., Levinson, D., Mudgett-Hunter, M., Tai, M. S., Novotny, J., Margolies, M. N., Ridge, R. J., Bruccoleri, R. E., Haber, E. and Crea, R. 1988. Protein engineering of antibody binding sites: recovery of specific activity in an anti-digoxin single-chain Fv analogue produced in Escherichia coli. Proc. Natl. Acad. Sci. USA. 85, 5879-5883.   DOI
9 Kalandadze, A., Galleno, M., Foncerrada, L., Strominger, J. L. and Wucherpfennig, K. W. 1996. Expression of recombinant HLA-DR2 molecules replacement of the hydrophobic transmembrane region by a leucine zipper dimerization motif allows the assembly and secretion of soluble DR αβ heterodimers. J. Biol. Chem. 271, 20156-20162.   DOI
10 Kim, C. H., Han, S. H., Kim, H. M., Han, J. Y., Lim, M. W. and Kim, J. K. 2015. The development of murine recombinant single-chain variable domain fragment (ScFv) specific to acute non-lymphocytic leukemia (ANLL) cell line HL60. Kor. J. Microbiol. 51, 115-125.   DOI
11 Scott, C. A., Garcia, K. C., Carbone, F. R., Wilson, I. A. and Teyton, L. 1996. Role of chain pairing for the production of functional soluble IA major histocompatibility complex class II molecules. J. Exp. Med. 183, 2087-2095.   DOI
12 Park, D. W., Kim, E. D., Kim, S. H., Han, J. Y. and Kim, J. K. 2011. The development of imerized Chicken Recombinant Single-chain Fv (ScFv) antibody using Leucine Zipper Motif. Kor. J. Microbiol. 47, 328-334.
13 Park, K. J., Park, D. W., Kim, C. H., Han, B. K., Park, T. S., Han, J. Y., Lillehoj, H. S. and Kim, J. K. 2005. Development and characterization of a recombinant chicken single-chain Fv antibody detecting Eimeria acervulina sporozoite antigen. Biotechnol. Lett. 27, 289-295.   DOI
14 Rau, R. 2002. Adalimumab (a fully human anti-tumour necrosis factor α monoclonal antibody) in the treatment of active rheumatoid arthritis: the initial results of five trials. Ann. Rheum. Dis. 61, ii70-ii73.   DOI
15 Reiter, Y., Brinkmann, U., Jung, S. H., Pastan, I. and Lee, B. 1995. Disulfide stabilization of antibody Fv: computer predictions and experimental evaluation. Protein Eng. 8, 1323-1331.   DOI
16 Risley, S., Thomas, M. A. and Bray, V. 2004. Rheumatoid arthritis, new standards of care: nursing implications of infliximab. J. Orthop. Nurs. 8, 41-49.   DOI
17 Shiraishi, T., Suzuyama, K., Okamoto, H., Mineta, T., Tabuchi, K., Nakayama, K., Shimizu, Y., Tohma, J., Ogihara, T., Naba, H., Mochizuki, H. and Nagata, S. 2004. Increased cytotoxicity of soluble Fas ligand by fusing isoleucine zipper motif. Biochem. Biophys. Res. Commun. 322, 197-202.   DOI
18 Skerra, A. and Pluckthun, A. 1988. Assembly of a functional immunoglobulin Fv fragment in Escherichia coli. Science 240, 1038-1041.   DOI
19 Tracey, D., Klareskog, L., Sasso, E. H., Salfeld, J. G. and Tak, P. P. 2008. Tumor necrosis factor antagonist mechanisms of action: A comprehensive review. Pharmacol. Ther. 117, 244-279.   DOI
20 Kim, S.W., Wu, S. and Kim, J. K. 2018. The development of anti-DR4 single-chain Fv (ScFv) antibody fused to Streptavidin. Kor. J. Microbiol. 54, 330-342.   DOI
21 Landschulz, W. H., Johnson, P. F. and McKnight, S. L. 1988. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science 240, 1759-1764.   DOI
22 Li, K., Zettlitz, K. A., Lipianskaya, J., Zhou, Y., Marks, J. D., Mallick, P., Reiter, R. E. and Wu, A. M. 2015. A fully human scFv phage display library for rapid antibody fragment reformatting. Protein Eng. Des. Sel. 28, 307-315   DOI
23 O'Shea, E. K., Rutkowski, R. and Kim, P. S. 1989. Evidence that the leucine zipper is a coiled coil. Science 243, 538-542.   DOI
24 Martin, A. C. and Thornton, J. M. 1996. Structural families in loops of homologous proteins: automatic classification, modelling and application to antibodies. J. Mol. Biol. 263, 800-815.   DOI
25 Min, W., Kim, J. K., Lillehoj, H. S ., Sohn, E. J., Han, J. Y., Song, K. D. and Lillehoj, E. P. 2001. Characterization of recombinant scFv antibody reactive with an apical antigen of Eimeria acervulina. Biotechnol. Lett. 23, 949-955.   DOI
26 O'Shea, E. K., Klemm, J. D., Kim, P. S. and Alber, T. 1991. X-ray structure of the GCN4 leucine zipper, a two-stranded, parallel coiled coil. Science 254, 539-544.   DOI
27 Pack, P., Kujau, M., Schroeckh, V., Knüpfer, U., Wenderoth, R., Riesenberg, D. and Plückthun, A. 1993. Improved bivalent miniantibodies, with identical avidity as whole antibodies, produced by high cell density fermentation of Escherichia coli. Biotechnology 11, 1271-1277.
28 Pack, P. and Plueckthun, A. 1992. Miniantibodies: use of amphipathic helixes to produce functional, flexibly linked dimeric FV fragments with high avidity in Escherichia coli. Biochemistry 31, 1579-1584.   DOI
29 Vinson, C., Acharya, A. and Taparowsky, E. J. 2006. Deciphering B-ZIP transcription factor interactions in vitro and in vivo. Biochim. Biophys. Acta Bioenerg. Gene Struct. Exp. 1759, 4-12.
30 Verma, R., Boleti, E. and George, A. 1998. Antibody engineering: comparison of bacterial, yeast, insect and mammalian expression systems. J. Immunol. Methods 216, 165-181.   DOI
31 Zimmermann, J., Voss, H., Schwager, C., Stegemann, J. and Ansorge, W. 1988. Automated Sanger dideoxy sequencing reaction protocol. FEBS Lett. 233, 432-436.   DOI
32 Bird, R. E., Hardman, K. D., Jacobson, J. W., Johnson, S., Kaufman, B. M., Lee, S. M., Lee, T., Pope, S. H., Riordan, G. S. and Whitlow, M. 1988. Single-chain antigen-binding proteins. Science 242, 423-426.   DOI