Heterologous overproduction of β-fructofuranosidase from yeast Xanthophyllomyces dendrorhous , an enzyme producing prebiotic sugars
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| 024 | 7 | 0 | |a 10.1007/s00253-014-6145-1 |2 doi |
| 035 | |a (NATIONALLICENCE)springer-10.1007/s00253-014-6145-1 | ||
| 245 | 0 | 0 | |a Heterologous overproduction of β-fructofuranosidase from yeast Xanthophyllomyces dendrorhous , an enzyme producing prebiotic sugars |h [Elektronische Daten] |c [María Gimeno-Pérez, Dolores Linde, Lucía Fernández-Arrojo, Francisco Plou, María Fernández-Lobato] |
| 520 | 3 | |a The β-fructofuranosidase Xd-INV from the yeast Xanthophyllomyces dendrorhous is the largest microbial enzyme producing neo-fructooligosaccharides (neo-FOS) known to date. It mainly synthesizes neokestose and neonystose, oligosaccharides with potentially improved prebiotic properties. The Xd-INV gene comprises an open reading frame of 1995bp, which encodes a 665-amino acid protein. Initial N-terminal sequencing of Xd-INV pointed to a majority extracellular protein of 595 amino acids lacking the first 70 residues (potential signal peptide). Functionality of the last 1785bp of Xd-INV gene was previously proved in Saccharomyces cerevisiae but only weak β-fructofuranosidase activity was quantified. In this study, different strategies to improve this enzyme level in a heterologous system have been used. Curiously, best results were obtained by increasing the protein N-terminus sequence in 39 amino acids, protein of 634 residues. The higher β-fructofuranosidase activity detected in this study, about 15U/mL, was obtained using Pichia pastoris and represents an improvement of about 1500 times the level previously obtained in a heterologous organism and doubles the best level of activity obtained by the natural producer. Heterologously expressed protein was purified and characterized biochemically and kinetically. Except by its glycosylation degree (10% lower) and thermal stability (4-5°C lower in the 60-85°C range), the properties of the heterologous enzyme, including ability to produce neo-FOS, remained unchanged. Interestingly, besides the neo-FOS referred before blastose was also detected (8-22g/L) in the reaction mixtures, making Xd-INV the first yeast enzyme producing this non-conventional disaccharide reported to date. | |
| 540 | |a Springer-Verlag Berlin Heidelberg, 2014 | ||
| 690 | 7 | |a Extracellular β-fructofuranosidase |2 nationallicence | |
| 690 | 7 | |a Xanthophyllomyces dendrorhous |2 nationallicence | |
| 690 | 7 | |a Neo-fructooligosaccharides |2 nationallicence | |
| 690 | 7 | |a Heterologous expression |2 nationallicence | |
| 690 | 7 | |a Pichia pastoris |2 nationallicence | |
| 690 | 7 | |a Blastose |2 nationallicence | |
| 700 | 1 | |a Gimeno-Pérez |D María |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | |
| 700 | 1 | |a Linde |D Dolores |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | |
| 700 | 1 | |a Fernández-Arrojo |D Lucía |u Instituto de Catálisis y Petroleoquímica, CSIC, Marie Curie 2, 28049, Madrid, Spain |4 aut | |
| 700 | 1 | |a Plou |D Francisco |u Instituto de Catálisis y Petroleoquímica, CSIC, Marie Curie 2, 28049, Madrid, Spain |4 aut | |
| 700 | 1 | |a Fernández-Lobato |D María |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | |
| 773 | 0 | |t Applied Microbiology and Biotechnology |d Springer Berlin Heidelberg |g 99/8(2015-04-01), 3459-3467 |x 0175-7598 |q 99:8<3459 |1 2015 |2 99 |o 253 | |
| 856 | 4 | 0 | |u https://doi.org/10.1007/s00253-014-6145-1 |q text/html |z Onlinezugriff via DOI |
| 898 | |a BK010053 |b XK010053 |c XK010000 | ||
| 900 | 7 | |a Metadata rights reserved |b Springer special CC-BY-NC licence |2 nationallicence | |
| 908 | |D 1 |a research-article |2 jats | ||
| 949 | |B NATIONALLICENCE |F NATIONALLICENCE |b NL-springer | ||
| 950 | |B NATIONALLICENCE |P 856 |E 40 |u https://doi.org/10.1007/s00253-014-6145-1 |q text/html |z Onlinezugriff via DOI | ||
| 950 | |B NATIONALLICENCE |P 700 |E 1- |a Gimeno-Pérez |D María |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | ||
| 950 | |B NATIONALLICENCE |P 700 |E 1- |a Linde |D Dolores |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | ||
| 950 | |B NATIONALLICENCE |P 700 |E 1- |a Fernández-Arrojo |D Lucía |u Instituto de Catálisis y Petroleoquímica, CSIC, Marie Curie 2, 28049, Madrid, Spain |4 aut | ||
| 950 | |B NATIONALLICENCE |P 700 |E 1- |a Plou |D Francisco |u Instituto de Catálisis y Petroleoquímica, CSIC, Marie Curie 2, 28049, Madrid, Spain |4 aut | ||
| 950 | |B NATIONALLICENCE |P 700 |E 1- |a Fernández-Lobato |D María |u Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, 28049, Madrid, Spain |4 aut | ||
| 950 | |B NATIONALLICENCE |P 773 |E 0- |t Applied Microbiology and Biotechnology |d Springer Berlin Heidelberg |g 99/8(2015-04-01), 3459-3467 |x 0175-7598 |q 99:8<3459 |1 2015 |2 99 |o 253 | ||