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Expression of exo-inulinase gene from Aspergillus niger 12 in E. coli strain Rosetta-gami B (DE3) and its characterization.
Biotechnol Prog. 2016 05; 32(3):629-37.BP

Abstract

Inulin is a linear carbohydrate polymer of fructose subunits (2-60) with terminal glucose units, produced as carbon storage in selected plants. It cannot directly be taken up by most microorganisms due to its large size, unless prior hydrolysis through inulinase enzymes occurs. The hydrolyzed inulin can be taken up by microbes and/or recovered and used industrially for the production of high fructose syrup, inulo-oligosaccharides, biofuel, and nutraceuticals. Cell-free enzymatic hydrolysis would be desirable for industrial applications, hence the recombinant expression, purification and characterization of an Aspergillus niger derived exo-inulinase was investigated in this study. The eukaroyototic exo-inulinase of Aspergillus niger 12 has been expressed, for the first time, in an E. coli strain [Rosetta-gami B (DE3)]. The molecular weight of recombinant exo-inulinase was estimated to be ∼81 kDa. The values of Km and Vmax of the recombinant exo-inulinase toward inulin were 5.3 ± 1.1 mM and 402.1 ± 53.1 µmol min(-1) mg(-1) protein, respectively. Towards sucrose the corresponding values were 12.20 ± 1.6 mM and 902.8 ± 40.2 µmol min(-1) mg(-1) protein towards sucrose. The S/I ratio was 2.24 ± 0.7, which is in the range of native inulinase. The optimum temperature and pH of the recombinant exo-inulinase towards inulin was 55°C and 5.0, while they were 50°C and 5.5 towards sucrose. The recombinant exo-inulinase activity towards inulin was enhanced by Cu(2+) and reduced by Fe(2+) , while its activity towards sucrose was enhanced by Co(2+) and reduced by Zn(2+) . © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:629-637, 2016.

Authors+Show Affiliations

Dept. of Chemical and Biochemical Engineering, University of Western Ontario, London, Ontario, N6A 5B9, Canada.Dept. of Chemical and Biochemical Engineering, Faculty of Engineering, University of Western Ontario, London, Ontario, N6A 5B9, Canada.Dept. of Chemical and Biochemical Engineering, Faculty of Engineering, University of Western Ontario, London, Ontario, N6A 5B9, Canada.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

26833959

Citation

Yedahalli, Shreyas S., et al. "Expression of Exo-inulinase Gene From Aspergillus Niger 12 in E. Coli Strain Rosetta-gami B (DE3) and Its Characterization." Biotechnology Progress, vol. 32, no. 3, 2016, pp. 629-37.
Yedahalli SS, Rehmann L, Bassi A. Expression of exo-inulinase gene from Aspergillus niger 12 in E. coli strain Rosetta-gami B (DE3) and its characterization. Biotechnol Prog. 2016;32(3):629-37.
Yedahalli, S. S., Rehmann, L., & Bassi, A. (2016). Expression of exo-inulinase gene from Aspergillus niger 12 in E. coli strain Rosetta-gami B (DE3) and its characterization. Biotechnology Progress, 32(3), 629-37. https://doi.org/10.1002/btpr.2238
Yedahalli SS, Rehmann L, Bassi A. Expression of Exo-inulinase Gene From Aspergillus Niger 12 in E. Coli Strain Rosetta-gami B (DE3) and Its Characterization. Biotechnol Prog. 2016;32(3):629-37. PubMed PMID: 26833959.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Expression of exo-inulinase gene from Aspergillus niger 12 in E. coli strain Rosetta-gami B (DE3) and its characterization. AU - Yedahalli,Shreyas S, AU - Rehmann,Lars, AU - Bassi,Amarjeet, Y1 - 2016/02/26/ PY - 2014/09/05/received PY - 2016/01/27/revised PY - 2016/2/3/entrez PY - 2016/2/3/pubmed PY - 2017/12/5/medline KW - Aspergillus niger 12 KW - E. coli KW - enzyme activity KW - exo-inulinase KW - inulin SP - 629 EP - 37 JF - Biotechnology progress JO - Biotechnol. Prog. VL - 32 IS - 3 N2 - Inulin is a linear carbohydrate polymer of fructose subunits (2-60) with terminal glucose units, produced as carbon storage in selected plants. It cannot directly be taken up by most microorganisms due to its large size, unless prior hydrolysis through inulinase enzymes occurs. The hydrolyzed inulin can be taken up by microbes and/or recovered and used industrially for the production of high fructose syrup, inulo-oligosaccharides, biofuel, and nutraceuticals. Cell-free enzymatic hydrolysis would be desirable for industrial applications, hence the recombinant expression, purification and characterization of an Aspergillus niger derived exo-inulinase was investigated in this study. The eukaroyototic exo-inulinase of Aspergillus niger 12 has been expressed, for the first time, in an E. coli strain [Rosetta-gami B (DE3)]. The molecular weight of recombinant exo-inulinase was estimated to be ∼81 kDa. The values of Km and Vmax of the recombinant exo-inulinase toward inulin were 5.3 ± 1.1 mM and 402.1 ± 53.1 µmol min(-1) mg(-1) protein, respectively. Towards sucrose the corresponding values were 12.20 ± 1.6 mM and 902.8 ± 40.2 µmol min(-1) mg(-1) protein towards sucrose. The S/I ratio was 2.24 ± 0.7, which is in the range of native inulinase. The optimum temperature and pH of the recombinant exo-inulinase towards inulin was 55°C and 5.0, while they were 50°C and 5.5 towards sucrose. The recombinant exo-inulinase activity towards inulin was enhanced by Cu(2+) and reduced by Fe(2+) , while its activity towards sucrose was enhanced by Co(2+) and reduced by Zn(2+) . © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:629-637, 2016. SN - 1520-6033 UR - https://www.unboundmedicine.com/medline/citation/26833959/Expression_of_exo_inulinase_gene_from_Aspergillus_niger_12_in_E__coli_strain_Rosetta_gami_B__DE3__and_its_characterization_ L2 - https://doi.org/10.1002/btpr.2238 DB - PRIME DP - Unbound Medicine ER -