Characterization of hyperbranched glycopolymers produced in vitro using enzymes

Autor: Xavier Roussel, Sandrine Laguerre, Florent Grimaud, Jean-Luc Putaux, Christophe D'Hulst, Anders Viksø-Nielsen, Gabrielle Potocki-Véronèse, Sophie Guilois, Christine Lancelon-Pin, Agnès Rolland-Sabaté, Alain Buléon
Přispěvatelé: Unité de recherche sur les Biopolymères, Interactions Assemblages (BIA), Institut National de la Recherche Agronomique (INRA), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Novozymes A/S, Agence Nationale de la Recherche [ANR-09-CP2D-07-01], inconnu, Inconnu, Centre de Recherches sur les Macromolécules Végétales (CERMAV), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Carret, Michèle
Jazyk: angličtina
Rok vydání: 2014
Předmět:
Sucrose
Hydrodynamic radius
Light
[SDV]Life Sciences [q-bio]
Dispersity
Asymmetrical flow field flow fractionation
02 engineering and technology
010402 general chemistry
Branching (polymer chemistry)
01 natural sciences
Biochemistry
Analytical Chemistry
Amylosucrase
Enzymatic hydrolysis
[SDV.IDA]Life Sciences [q-bio]/Food engineering
Glycogen branching enzyme
Scattering
Radiation
[SPI.GPROC]Engineering Sciences [physics]/Chemical and Process Engineering
Glucans
ComputingMilieux_MISCELLANEOUS
Chromatography
Molar mass
Bacteria
Molecular Structure
biology
Chemistry
Branching enzyme
021001 nanoscience & nanotechnology
Fractionation
Field Flow
0104 chemical sciences
Molecular Weight
Hyperbranched alpha-glucan
Glucosyltransferases
Amylases
biology.protein
Biophysics
Macromolecular size distribution
0210 nano-technology
Glycogen
Macromolecule
Zdroj: Analytical and Bioanalytical Chemistry
Analytical and Bioanalytical Chemistry, 2014, 406 (6), pp.1607-1618. ⟨10.1007/s00216-013-7403-2⟩
Analytical and Bioanalytical Chemistry, Springer Verlag, 2014, pp.1607-1618
Analytical and Bioanalytical Chemistry, Springer Verlag, 2014, 406 (6), pp.1607-1618. ⟨10.1007/s00216-013-7403-2⟩
ISSN: 1618-2642
1618-2650
DOI: 10.1007/s00216-013-7403-2⟩
Popis: International audience; Asymmetrical flow field flow fractionation (AF4) has proven to be a very powerful and quantitative method for the determination of the macromolecular structure of high molar mass branched biopolymers, when coupled with multi-angle laser light scattering (MALLS). This work describes a detailed investigation of the macromolecular structure of native glycogens and hyperbranched alpha-glucans (HBPs), with average molar mass ranging from 2 x 10(6) to 4.3 x 10(7) g mol(-1), which are not well fractionated by means of classical size-exclusion chromatography. HBPs were enzymatically produced from sucrose by the tandem action of an amylosucrase and a branching enzyme mimicking in vitro the elongation and branching steps involved in glycogen biosynthesis. Size and molar mass distributions were studied by AF4, coupled with online quasi-elastic light scattering (QELS) and transmission electron microscopy. AF4-MALLS-QELS has shown a remarkable agreement between hydrodynamic radii obtained by online QELS and by AF4 theory in normal mode with constant cross flow. Molar mass, size, and dispersity were shown to significantly increase with initial sucrose concentration, and to decrease when the branching enzyme activity increases. Several populations with different size range were observed: the amount of small size molecules decreasing with increasing sucrose concentration. The spherical and dense global conformation thus highlighted was partly similar to native glycogens. A more detailed study of HBPs synthesized from low and high initial sucrose concentrations was performed using complementary enzymatic hydrolysis of external chains and chromatography. It emphasized a more homogeneous branching pattern than native glycogens with a denser core and shorter external chains.
Databáze: OpenAIRE
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