Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.

Further improvement of the thermostability of inherently thermostable proteins is an attractive challenge because more thermostable proteins are industrially more useful and serve as better scaffolds for protein engineering. To establish guidelines that can be applied for the rational design of hype...

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Autores principales:	Satoshi Akanuma, Minako Yamaguchi, Akihiko Yamagishi
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/4754a09927444b30896cef2d4e0efe85
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spelling	oai:doaj.org-article:4754a09927444b30896cef2d4e0efe852021-12-02T20:16:39ZComprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.1932-620310.1371/journal.pone.0258821https://doaj.org/article/4754a09927444b30896cef2d4e0efe852021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0258821https://doaj.org/toc/1932-6203Further improvement of the thermostability of inherently thermostable proteins is an attractive challenge because more thermostable proteins are industrially more useful and serve as better scaffolds for protein engineering. To establish guidelines that can be applied for the rational design of hyperthermostable proteins, we compared the amino acid sequences of two ancestral nucleoside diphosphate kinases, Arc1 and Bac1, reconstructed in our previous study. Although Bac1 is a thermostable protein whose unfolding temperature is around 100°C, Arc1 is much more thermostable with an unfolding temperature of 114°C. However, only 12 out of 139 amino acids are different between the two sequences. In this study, one or a combination of amino acid(s) in Bac1 was/were substituted by a residue(s) found in Arc1 at the same position(s). The best mutant, which contained three amino acid substitutions (S108D, G116A and L120P substitutions), showed an unfolding temperature more than 10°C higher than that of Bac1. Furthermore, a combination of the other nine amino acid substitutions also led to improved thermostability of Bac1, although the effects of individual substitutions were small. Therefore, not only the sum of the contributions of individual amino acids, but also the synergistic effects of multiple amino acids are deeply involved in the stability of a hyperthermostable protein. Such insights will be helpful for future rational design of hyperthermostable proteins.Satoshi AkanumaMinako YamaguchiAkihiko YamagishiPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 10, p e0258821 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Satoshi Akanuma Minako Yamaguchi Akihiko Yamagishi Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
description	Further improvement of the thermostability of inherently thermostable proteins is an attractive challenge because more thermostable proteins are industrially more useful and serve as better scaffolds for protein engineering. To establish guidelines that can be applied for the rational design of hyperthermostable proteins, we compared the amino acid sequences of two ancestral nucleoside diphosphate kinases, Arc1 and Bac1, reconstructed in our previous study. Although Bac1 is a thermostable protein whose unfolding temperature is around 100°C, Arc1 is much more thermostable with an unfolding temperature of 114°C. However, only 12 out of 139 amino acids are different between the two sequences. In this study, one or a combination of amino acid(s) in Bac1 was/were substituted by a residue(s) found in Arc1 at the same position(s). The best mutant, which contained three amino acid substitutions (S108D, G116A and L120P substitutions), showed an unfolding temperature more than 10°C higher than that of Bac1. Furthermore, a combination of the other nine amino acid substitutions also led to improved thermostability of Bac1, although the effects of individual substitutions were small. Therefore, not only the sum of the contributions of individual amino acids, but also the synergistic effects of multiple amino acids are deeply involved in the stability of a hyperthermostable protein. Such insights will be helpful for future rational design of hyperthermostable proteins.
format	article
author	Satoshi Akanuma Minako Yamaguchi Akihiko Yamagishi
author_facet	Satoshi Akanuma Minako Yamaguchi Akihiko Yamagishi
author_sort	Satoshi Akanuma
title	Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
title_short	Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
title_full	Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
title_fullStr	Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
title_full_unstemmed	Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
title_sort	comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/4754a09927444b30896cef2d4e0efe85
work_keys_str_mv	AT satoshiakanuma comprehensivemutagenesistoidentifyaminoacidresiduescontributingtothedifferenceinthermostabilitybetweentwooriginallythermostableancestralproteins AT minakoyamaguchi comprehensivemutagenesistoidentifyaminoacidresiduescontributingtothedifferenceinthermostabilitybetweentwooriginallythermostableancestralproteins AT akihikoyamagishi comprehensivemutagenesistoidentifyaminoacidresiduescontributingtothedifferenceinthermostabilitybetweentwooriginallythermostableancestralproteins
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Comprehensive mutagenesis to identify amino acid residues contributing to the difference in thermostability between two originally thermostable ancestral proteins.

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