Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.

This paper reports rational engineering of Trypanosoma rangeli sialidase to develop an effective enzyme for a potentially important type of reactivity: production of sialylated prebiotic glycans. The Trypanosoma cruzi trans-sialidase and the homologous T. rangeli sialidase has previously been used t...

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Autores principales:	Carsten Jers, Malwina Michalak, Dorte M Larsen, Kasper P Kepp, Haiying Li, Yao Guo, Finn Kirpekar, Anne S Meyer, Jørn D Mikkelsen
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2014
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/7ce5c01b05ce4c2b82b60c94e2281430
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spelling	oai:doaj.org-article:7ce5c01b05ce4c2b82b60c94e22814302021-11-18T08:39:00ZRational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.1932-620310.1371/journal.pone.0083902https://doaj.org/article/7ce5c01b05ce4c2b82b60c94e22814302014-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24404142/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203This paper reports rational engineering of Trypanosoma rangeli sialidase to develop an effective enzyme for a potentially important type of reactivity: production of sialylated prebiotic glycans. The Trypanosoma cruzi trans-sialidase and the homologous T. rangeli sialidase has previously been used to investigate the structural requirements for trans-sialidase activity. We observed that the T. cruzi trans-sialidase has a seven-amino-acid motif (197-203) at the border of the substrate binding cleft. The motif differs substantially in chemical properties and substitution probability from the homologous sialidase, and we hypothesised that this motif is important for trans-sialidase activity. The 197-203 motif is strongly positively charged with a marked change in hydrogen bond donor capacity as compared to the sialidase. To investigate the role of this motif, we expressed and characterised a T. rangeli sialidase mutant, Tr13. Conditions for efficient trans-sialylation were determined, and Tr13's acceptor specificity demonstrated promiscuity with respect to the acceptor molecule enabling sialylation of glycans containing terminal galactose and glucose and even monomers of glucose and fucose. Sialic acid is important in association with human milk oligosaccharides, and Tr13 was shown to sialylate a number of established and potential prebiotics. Initial evaluation of prebiotic potential using pure cultures demonstrated, albeit not selectively, growth of Bifidobacteria. Since the 197-203 motif stands out in the native trans-sialidase, is markedly different from the wild-type sialidase compared to previous mutants, and is shown here to confer efficient and broad trans-sialidase activity, we suggest that this motif can serve as a framework for future optimization of trans-sialylation towards prebiotic production.Carsten JersMalwina MichalakDorte M LarsenKasper P KeppHaiying LiYao GuoFinn KirpekarAnne S MeyerJørn D MikkelsenPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 9, Iss 1, p e83902 (2014)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Carsten Jers Malwina Michalak Dorte M Larsen Kasper P Kepp Haiying Li Yao Guo Finn Kirpekar Anne S Meyer Jørn D Mikkelsen Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
description	This paper reports rational engineering of Trypanosoma rangeli sialidase to develop an effective enzyme for a potentially important type of reactivity: production of sialylated prebiotic glycans. The Trypanosoma cruzi trans-sialidase and the homologous T. rangeli sialidase has previously been used to investigate the structural requirements for trans-sialidase activity. We observed that the T. cruzi trans-sialidase has a seven-amino-acid motif (197-203) at the border of the substrate binding cleft. The motif differs substantially in chemical properties and substitution probability from the homologous sialidase, and we hypothesised that this motif is important for trans-sialidase activity. The 197-203 motif is strongly positively charged with a marked change in hydrogen bond donor capacity as compared to the sialidase. To investigate the role of this motif, we expressed and characterised a T. rangeli sialidase mutant, Tr13. Conditions for efficient trans-sialylation were determined, and Tr13's acceptor specificity demonstrated promiscuity with respect to the acceptor molecule enabling sialylation of glycans containing terminal galactose and glucose and even monomers of glucose and fucose. Sialic acid is important in association with human milk oligosaccharides, and Tr13 was shown to sialylate a number of established and potential prebiotics. Initial evaluation of prebiotic potential using pure cultures demonstrated, albeit not selectively, growth of Bifidobacteria. Since the 197-203 motif stands out in the native trans-sialidase, is markedly different from the wild-type sialidase compared to previous mutants, and is shown here to confer efficient and broad trans-sialidase activity, we suggest that this motif can serve as a framework for future optimization of trans-sialylation towards prebiotic production.
format	article
author	Carsten Jers Malwina Michalak Dorte M Larsen Kasper P Kepp Haiying Li Yao Guo Finn Kirpekar Anne S Meyer Jørn D Mikkelsen
author_facet	Carsten Jers Malwina Michalak Dorte M Larsen Kasper P Kepp Haiying Li Yao Guo Finn Kirpekar Anne S Meyer Jørn D Mikkelsen
author_sort	Carsten Jers
title	Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
title_short	Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
title_full	Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
title_fullStr	Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
title_full_unstemmed	Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
title_sort	rational design of a new trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.
publisher	Public Library of Science (PLoS)
publishDate	2014
url	https://doaj.org/article/7ce5c01b05ce4c2b82b60c94e2281430
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Rational design of a new Trypanosoma rangeli trans-sialidase for efficient sialylation of glycans.

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