Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities

Abstract Single domain antibodies (sdAbs), made of natural single variable regions of camelid or cartilaginous fish antibodies, or unpaired variable regions of mouse or human IgGs, are some of the more promising biologic modalities. However, such conventional sdAbs have difficulties of either using...

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Autores principales:	Naoya Shinozaki, Ryuji Hashimoto, Kiichi Fukui, Susumu Uchiyama
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/ad22077988eb44aeab1bd0872d14680f
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spelling	oai:doaj.org-article:ad22077988eb44aeab1bd0872d14680f2021-12-02T11:40:13ZEfficient generation of single domain antibodies with high affinities and enhanced thermal stabilities10.1038/s41598-017-06277-x2045-2322https://doaj.org/article/ad22077988eb44aeab1bd0872d14680f2017-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-06277-xhttps://doaj.org/toc/2045-2322Abstract Single domain antibodies (sdAbs), made of natural single variable regions of camelid or cartilaginous fish antibodies, or unpaired variable regions of mouse or human IgGs, are some of the more promising biologic modalities. However, such conventional sdAbs have difficulties of either using unwieldy animals for immunization or having high affinity deficiencies. Herein, we offer a versatile method to generate rabbit variable domain of heavy chain (rVH) derived sdAbs with high affinities (K D values of single digit nM or less) and enhanced thermal stabilities (equal to or even higher than those of camelid derived sdAbs). It was found that a variety of rVH binders, including those with high affinities, were efficiently acquired using an rVH-displaying phage library produced at a low temperature of 16 °C. By a simple method to introduce an additional disulfide bond, their unfolding temperatures were increased by more than 20 °C without severe loss of binding affinity. Differential scanning calorimetry analysis suggested that this highly efficient thermal stabilization was mainly attributed to the entropic contribution and unique thermodynamic character of the rVHs.Naoya ShinozakiRyuji HashimotoKiichi FukuiSusumu UchiyamaNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-11 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Naoya Shinozaki Ryuji Hashimoto Kiichi Fukui Susumu Uchiyama Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
description	Abstract Single domain antibodies (sdAbs), made of natural single variable regions of camelid or cartilaginous fish antibodies, or unpaired variable regions of mouse or human IgGs, are some of the more promising biologic modalities. However, such conventional sdAbs have difficulties of either using unwieldy animals for immunization or having high affinity deficiencies. Herein, we offer a versatile method to generate rabbit variable domain of heavy chain (rVH) derived sdAbs with high affinities (K D values of single digit nM or less) and enhanced thermal stabilities (equal to or even higher than those of camelid derived sdAbs). It was found that a variety of rVH binders, including those with high affinities, were efficiently acquired using an rVH-displaying phage library produced at a low temperature of 16 °C. By a simple method to introduce an additional disulfide bond, their unfolding temperatures were increased by more than 20 °C without severe loss of binding affinity. Differential scanning calorimetry analysis suggested that this highly efficient thermal stabilization was mainly attributed to the entropic contribution and unique thermodynamic character of the rVHs.
format	article
author	Naoya Shinozaki Ryuji Hashimoto Kiichi Fukui Susumu Uchiyama
author_facet	Naoya Shinozaki Ryuji Hashimoto Kiichi Fukui Susumu Uchiyama
author_sort	Naoya Shinozaki
title	Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
title_short	Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
title_full	Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
title_fullStr	Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
title_full_unstemmed	Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
title_sort	efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/ad22077988eb44aeab1bd0872d14680f
work_keys_str_mv	AT naoyashinozaki efficientgenerationofsingledomainantibodieswithhighaffinitiesandenhancedthermalstabilities AT ryujihashimoto efficientgenerationofsingledomainantibodieswithhighaffinitiesandenhancedthermalstabilities AT kiichifukui efficientgenerationofsingledomainantibodieswithhighaffinitiesandenhancedthermalstabilities AT susumuuchiyama efficientgenerationofsingledomainantibodieswithhighaffinitiesandenhancedthermalstabilities
_version_	1718395719479459840

Efficient generation of single domain antibodies with high affinities and enhanced thermal stabilities

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