Immobilization of β-galactosidase and α-mannosidase onto magnetic nanoparticles: A strategy for increasing the potentiality of valuable glycomic tools for glycosylation analysis and biological role determination of glycoconjugates

Ernesto Rodríguez, Karen Francia, Natalie Brossard, Juan J García Vallejo, Hakan Kalay, Yvette van Kooyk, Teresa Freire, Cecilia Giacomini

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Glycans present in biological glycoconjugates have several structural and functional roles. Elucidation of glycan structure and biological function is critical to understand their role in physiological and pathogenic process, enabling the development of diagnostic methods and disease treatment. Immobilized glycosidases are powerful tools for glycan analysis, as they are able to remove specific carbohydrates without altering the protein structure. Here we describe the individual immobilization of Aspergillus oryzae β-galactosidase and Canavalia ensiformis α-mannosidase onto agarose and silica magnetic nanoparticles activated with cyanate ester groups. High immobilization yields (70-90%) were achieved, keeping above 60% of its original activity. Immobilized glycosidases were effective in the selective deglycosylation of model glycoproteins and a Fasciola hepatica lysate, evidenced by a decrease in specific lectin recognition of 40-50% after enzymatic deglycosylation. Immobilized glycosidases were reused for several deglycosylation cycles without loss of effectiveness. Their use was extended to the elucidation of the glycan role of native glycoconjugates. A decrease in the recognition of lactoferrin treated with α-mannosidase by a C-type lectin receptor, DC-SIGN was found. Also the specific deglycosylation of a F. hepatica lysate demonstrated the relevance of mannosylated glycans in the induction of Th2/Treg immune responses during the infection. Our results show successful immobilization of specific glycosidases in nano-supports and validate their utility to identify glycans biological functions.

Original languageEnglish
Pages (from-to)45-55
Number of pages11
JournalEnzyme and microbial technology
Volume117
DOIs
Publication statusPublished - Oct 2018

Cite this

@article{7b3ec010ba524a538768026f0dfdf33a,
title = "Immobilization of β-galactosidase and α-mannosidase onto magnetic nanoparticles: A strategy for increasing the potentiality of valuable glycomic tools for glycosylation analysis and biological role determination of glycoconjugates",
abstract = "Glycans present in biological glycoconjugates have several structural and functional roles. Elucidation of glycan structure and biological function is critical to understand their role in physiological and pathogenic process, enabling the development of diagnostic methods and disease treatment. Immobilized glycosidases are powerful tools for glycan analysis, as they are able to remove specific carbohydrates without altering the protein structure. Here we describe the individual immobilization of Aspergillus oryzae β-galactosidase and Canavalia ensiformis α-mannosidase onto agarose and silica magnetic nanoparticles activated with cyanate ester groups. High immobilization yields (70-90{\%}) were achieved, keeping above 60{\%} of its original activity. Immobilized glycosidases were effective in the selective deglycosylation of model glycoproteins and a Fasciola hepatica lysate, evidenced by a decrease in specific lectin recognition of 40-50{\%} after enzymatic deglycosylation. Immobilized glycosidases were reused for several deglycosylation cycles without loss of effectiveness. Their use was extended to the elucidation of the glycan role of native glycoconjugates. A decrease in the recognition of lactoferrin treated with α-mannosidase by a C-type lectin receptor, DC-SIGN was found. Also the specific deglycosylation of a F. hepatica lysate demonstrated the relevance of mannosylated glycans in the induction of Th2/Treg immune responses during the infection. Our results show successful immobilization of specific glycosidases in nano-supports and validate their utility to identify glycans biological functions.",
author = "Ernesto Rodr{\'i}guez and Karen Francia and Natalie Brossard and {Garc{\'i}a Vallejo}, {Juan J} and Hakan Kalay and {van Kooyk}, Yvette and Teresa Freire and Cecilia Giacomini",
note = "Copyright {\circledC} 2018 Elsevier Inc. All rights reserved.",
year = "2018",
month = "10",
doi = "10.1016/j.enzmictec.2018.05.012",
language = "English",
volume = "117",
pages = "45--55",
journal = "Enzyme and microbial technology",
issn = "0141-0229",
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TY - JOUR

T1 - Immobilization of β-galactosidase and α-mannosidase onto magnetic nanoparticles

T2 - A strategy for increasing the potentiality of valuable glycomic tools for glycosylation analysis and biological role determination of glycoconjugates

AU - Rodríguez, Ernesto

AU - Francia, Karen

AU - Brossard, Natalie

AU - García Vallejo, Juan J

AU - Kalay, Hakan

AU - van Kooyk, Yvette

AU - Freire, Teresa

AU - Giacomini, Cecilia

N1 - Copyright © 2018 Elsevier Inc. All rights reserved.

PY - 2018/10

Y1 - 2018/10

N2 - Glycans present in biological glycoconjugates have several structural and functional roles. Elucidation of glycan structure and biological function is critical to understand their role in physiological and pathogenic process, enabling the development of diagnostic methods and disease treatment. Immobilized glycosidases are powerful tools for glycan analysis, as they are able to remove specific carbohydrates without altering the protein structure. Here we describe the individual immobilization of Aspergillus oryzae β-galactosidase and Canavalia ensiformis α-mannosidase onto agarose and silica magnetic nanoparticles activated with cyanate ester groups. High immobilization yields (70-90%) were achieved, keeping above 60% of its original activity. Immobilized glycosidases were effective in the selective deglycosylation of model glycoproteins and a Fasciola hepatica lysate, evidenced by a decrease in specific lectin recognition of 40-50% after enzymatic deglycosylation. Immobilized glycosidases were reused for several deglycosylation cycles without loss of effectiveness. Their use was extended to the elucidation of the glycan role of native glycoconjugates. A decrease in the recognition of lactoferrin treated with α-mannosidase by a C-type lectin receptor, DC-SIGN was found. Also the specific deglycosylation of a F. hepatica lysate demonstrated the relevance of mannosylated glycans in the induction of Th2/Treg immune responses during the infection. Our results show successful immobilization of specific glycosidases in nano-supports and validate their utility to identify glycans biological functions.

AB - Glycans present in biological glycoconjugates have several structural and functional roles. Elucidation of glycan structure and biological function is critical to understand their role in physiological and pathogenic process, enabling the development of diagnostic methods and disease treatment. Immobilized glycosidases are powerful tools for glycan analysis, as they are able to remove specific carbohydrates without altering the protein structure. Here we describe the individual immobilization of Aspergillus oryzae β-galactosidase and Canavalia ensiformis α-mannosidase onto agarose and silica magnetic nanoparticles activated with cyanate ester groups. High immobilization yields (70-90%) were achieved, keeping above 60% of its original activity. Immobilized glycosidases were effective in the selective deglycosylation of model glycoproteins and a Fasciola hepatica lysate, evidenced by a decrease in specific lectin recognition of 40-50% after enzymatic deglycosylation. Immobilized glycosidases were reused for several deglycosylation cycles without loss of effectiveness. Their use was extended to the elucidation of the glycan role of native glycoconjugates. A decrease in the recognition of lactoferrin treated with α-mannosidase by a C-type lectin receptor, DC-SIGN was found. Also the specific deglycosylation of a F. hepatica lysate demonstrated the relevance of mannosylated glycans in the induction of Th2/Treg immune responses during the infection. Our results show successful immobilization of specific glycosidases in nano-supports and validate their utility to identify glycans biological functions.

U2 - 10.1016/j.enzmictec.2018.05.012

DO - 10.1016/j.enzmictec.2018.05.012

M3 - Article

VL - 117

SP - 45

EP - 55

JO - Enzyme and microbial technology

JF - Enzyme and microbial technology

SN - 0141-0229

ER -