Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac

Abstract Pseudaminic acids present on the surface of pathogenic bacteria, including gut pathogens Campylobacter jejuni and Helicobacter pylori, are postulated to play influential roles in the etiology of associated infectious diseases through modulating flagella assembly and recognition of bacteria...

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Autores principales:	Harriet S. Chidwick, Emily K. P. Flack, Tessa Keenan, Julia Walton, Gavin H. Thomas, Martin A. Fascione
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/748cb75648664ed68c7f0a76f00b64a8
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spelling	oai:doaj.org-article:748cb75648664ed68c7f0a76f00b64a82021-12-02T11:37:26ZReconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac10.1038/s41598-021-83707-x2045-2322https://doaj.org/article/748cb75648664ed68c7f0a76f00b64a82021-02-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-83707-xhttps://doaj.org/toc/2045-2322Abstract Pseudaminic acids present on the surface of pathogenic bacteria, including gut pathogens Campylobacter jejuni and Helicobacter pylori, are postulated to play influential roles in the etiology of associated infectious diseases through modulating flagella assembly and recognition of bacteria by the human immune system. Yet they are underexplored compared to other areas of glycoscience, in particular enzymes responsible for the glycosyltransfer of these sugars in bacteria are still to be unambiguously characterised. This can be largely attributed to a lack of access to nucleotide-activated pseudaminic acid glycosyl donors, such as CMP-Pse5Ac7Ac. Herein we reconstitute the biosynthesis of Pse5Ac7Ac in vitro using enzymes from C. jejuni (PseBCHGI) in the process optimising coupled turnover with PseBC using deuterium wash in experiments, and establishing a method for co-factor regeneration in PseH tunover. Furthermore we establish conditions for purification of a soluble CMP-Pse5Ac7Ac synthetase enzyme PseF from Aeromonas caviae and utilise it in combination with the C. jejuni enzymes to achieve practical preparative synthesis of CMP-Pse5Ac7Ac in vitro, facilitating future biological studies.Harriet S. ChidwickEmily K. P. FlackTessa KeenanJulia WaltonGavin H. ThomasMartin A. FascioneNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Harriet S. Chidwick Emily K. P. Flack Tessa Keenan Julia Walton Gavin H. Thomas Martin A. Fascione Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
description	Abstract Pseudaminic acids present on the surface of pathogenic bacteria, including gut pathogens Campylobacter jejuni and Helicobacter pylori, are postulated to play influential roles in the etiology of associated infectious diseases through modulating flagella assembly and recognition of bacteria by the human immune system. Yet they are underexplored compared to other areas of glycoscience, in particular enzymes responsible for the glycosyltransfer of these sugars in bacteria are still to be unambiguously characterised. This can be largely attributed to a lack of access to nucleotide-activated pseudaminic acid glycosyl donors, such as CMP-Pse5Ac7Ac. Herein we reconstitute the biosynthesis of Pse5Ac7Ac in vitro using enzymes from C. jejuni (PseBCHGI) in the process optimising coupled turnover with PseBC using deuterium wash in experiments, and establishing a method for co-factor regeneration in PseH tunover. Furthermore we establish conditions for purification of a soluble CMP-Pse5Ac7Ac synthetase enzyme PseF from Aeromonas caviae and utilise it in combination with the C. jejuni enzymes to achieve practical preparative synthesis of CMP-Pse5Ac7Ac in vitro, facilitating future biological studies.
format	article
author	Harriet S. Chidwick Emily K. P. Flack Tessa Keenan Julia Walton Gavin H. Thomas Martin A. Fascione
author_facet	Harriet S. Chidwick Emily K. P. Flack Tessa Keenan Julia Walton Gavin H. Thomas Martin A. Fascione
author_sort	Harriet S. Chidwick
title	Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
title_short	Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
title_full	Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
title_fullStr	Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
title_full_unstemmed	Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac
title_sort	reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (pse5ac7ac) enables preparative enzymatic synthesis of cmp-pse5ac7ac
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/748cb75648664ed68c7f0a76f00b64a8
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Reconstitution and optimisation of the biosynthesis of bacterial sugar pseudaminic acid (Pse5Ac7Ac) enables preparative enzymatic synthesis of CMP-Pse5Ac7Ac

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