Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery

Abstract A fusion protein comprising an antibody and a cationic peptide, such as arginine-9 (R9), is a candidate molecule for efficient and cell-specific delivery of siRNA into cells in order to reduce the side effects of nucleic acid drugs. However, their expression in bacterial hosts, required for...

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Autores principales:	Yu Ando, Hikaru Nakazawa, Daisuke Miura, Maho Otake, Mitsuo Umetsu
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/a93f63012e644dfbadfe780137a79059
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spelling	oai:doaj.org-article:a93f63012e644dfbadfe780137a790592021-11-14T12:21:04ZEnzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery10.1038/s41598-021-01331-12045-2322https://doaj.org/article/a93f63012e644dfbadfe780137a790592021-11-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-01331-1https://doaj.org/toc/2045-2322Abstract A fusion protein comprising an antibody and a cationic peptide, such as arginine-9 (R9), is a candidate molecule for efficient and cell-specific delivery of siRNA into cells in order to reduce the side effects of nucleic acid drugs. However, their expression in bacterial hosts, required for their development, often fails, impeding research progress. In this study, we separately prepared anti-EGFR nanobodies with the K-tag sequence MRHKGS at the C-terminus and R9 with the Q-tag sequence LLQG at the N-terminus, and enzymatically ligated them in vitro by microbial transglutaminase to generate Nanobody-R9, which is not expressed as a fused protein in E. coli. Nanobody-R9 was synthesized at a maximum binding efficiency of 85.1%, without changing the binding affinity of the nanobody for the antigen. Nanobody-R9 successfully delivered siRNA into the cells, and the cellular influx of siRNA increased with increase in the ratio of Nanobody-R9 to siRNA. We further demonstrated that the Nanobody-R9–siRNA complex, at a 30:1 ratio, induced an approximately 58.6% reduction in the amount of target protein due to RNAi in mRNA compared to lipofectamine.Yu AndoHikaru NakazawaDaisuke MiuraMaho OtakeMitsuo UmetsuNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Yu Ando Hikaru Nakazawa Daisuke Miura Maho Otake Mitsuo Umetsu Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
description	Abstract A fusion protein comprising an antibody and a cationic peptide, such as arginine-9 (R9), is a candidate molecule for efficient and cell-specific delivery of siRNA into cells in order to reduce the side effects of nucleic acid drugs. However, their expression in bacterial hosts, required for their development, often fails, impeding research progress. In this study, we separately prepared anti-EGFR nanobodies with the K-tag sequence MRHKGS at the C-terminus and R9 with the Q-tag sequence LLQG at the N-terminus, and enzymatically ligated them in vitro by microbial transglutaminase to generate Nanobody-R9, which is not expressed as a fused protein in E. coli. Nanobody-R9 was synthesized at a maximum binding efficiency of 85.1%, without changing the binding affinity of the nanobody for the antigen. Nanobody-R9 successfully delivered siRNA into the cells, and the cellular influx of siRNA increased with increase in the ratio of Nanobody-R9 to siRNA. We further demonstrated that the Nanobody-R9–siRNA complex, at a 30:1 ratio, induced an approximately 58.6% reduction in the amount of target protein due to RNAi in mRNA compared to lipofectamine.
format	article
author	Yu Ando Hikaru Nakazawa Daisuke Miura Maho Otake Mitsuo Umetsu
author_facet	Yu Ando Hikaru Nakazawa Daisuke Miura Maho Otake Mitsuo Umetsu
author_sort	Yu Ando
title	Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
title_short	Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
title_full	Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
title_fullStr	Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
title_full_unstemmed	Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery
title_sort	enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific sirna delivery
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/a93f63012e644dfbadfe780137a79059
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Enzymatic ligation of an antibody and arginine 9 peptide for efficient and cell-specific siRNA delivery

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