Construction of Genetically Modified Lactococcus lactis Producing Anti-human-CTLA-4 Single-Chain Fragment Variable
| Autor: | Suguru Shigemori, Aito Murakami, Masami Tsukagoshi, Asami Ueda, Takashi Sato, Tasuku Ogita, Fu Namai, Takeshi Shimosato |
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| Rok vydání: | 2020 |
| Předmět: |
0106 biological sciences
Bioengineering Protein Engineering 01 natural sciences Applied Microbiology and Biotechnology Biochemistry law.invention 03 medical and health sciences Antineoplastic Agents Immunological Immune system Antigen law 010608 biotechnology Gene expression Humans CTLA-4 Antigen Cloning Molecular Promoter Regions Genetic Molecular Biology Nisin 030304 developmental biology 0303 health sciences biology Chemistry Lactococcus lactis Gene Expression Regulation Bacterial biology.organism_classification Molecular biology Recombinant Proteins Immune checkpoint Genetically modified organism biology.protein Recombinant DNA Antibody Single-Chain Antibodies Biotechnology |
| Zdroj: | Molecular Biotechnology. 62:572-579 |
| ISSN: | 1559-0305 1073-6085 |
| Popis: | Lactic acid bacteria are human commensal organisms that have immunomodulatory and metabolism-promoting effects. In addition, due to the increasing demand for biopharmaceuticals, genetically modified lactic acid bacteria (gmLAB) that produce recombinant proteins are expected to be used as microbial therapeutics and next-generation probiotics. In this study, we constructed a gmLAB strain that produces anti-human cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) single-chain fragment variable (CTLA4scFv) for possible use in a cancer treatment strategy using gmLAB. CTLA-4, an immune checkpoint molecule, suppresses the anti-cancer immune response; thus, inhibition of CTLA-4 signaling is important in cancer therapy. In this study, we designed a CTLA4scFv composed of a heavy and light chain of the variable region from anti-human CTLA-4 antibody connected by a flexible peptide linker. CTLA4scFv was expressed using nisin controlled gene expression (NICE) system, a lactococcal inducible gene expression system, and the DNA sequence encoding CTLA4scFv was inserted downstream of the PnisA promoter of the gene expression vector pNZ8148#2. Furthermore, expression of recombinant CTLA4scFv was confirmed by Western blotting, and the immunoreactivity of recombinant CTLA4scFv against human CTLA-4 protein was examined using ELISA. We speculate that gmLAB producing bioactive CTLA4scFv will become an attractive approach for cancer treatment. |
| Databáze: | OpenAIRE |
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