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Biotechnology Letters

, Volume 40, Issue 6, pp 949–955 | Cite as

The GlaA signal peptide substantially increases the expression and secretion of α-galactosidase in Aspergillus niger

  • Yue Xu
  • Yan-hui Wang
  • Tian-qi Liu
  • Hui Zhang
  • He Zhang
  • Jie Li
Original Research Paper
  • 193 Downloads

Abstract

Objective

α-Galactosidases are widely used in many fields. It is necessary to improve the production of enzymes through microbiological processes. The aim of this study was to construct recombinant Aspergillus niger strains with high α-galactosidase production.

Results

Two recombinant A. niger strains were constructed: AB and AGB. The recombinant AB strain contained the α-galactosidase aglB gene from A. niger with its native AglB signal peptide regulated by the glucoamylase promoter. In the AGB recombinant strain, the AglB signal peptide was replaced with the glucoamylase (GlaA) signal peptide. The extracellular maximum α-galactosidase activity of the AGB strain was 215.7 U/ml and that of the AB strain was 9.8 U/mL. The optimal conditions for α-galactosidase were pH 3.5 and 35 °C.

Conclusions

The GlaA signal peptide substantially increased the yield of secreted α-galactosidase in A. niger. This recombinant strain holds great potential for industrial applications.

Keywords

Aspergillus niger α-galactosidase Secretion Signal peptide 

Notes

Acknowledgements

This work was supported by the Special Scientific Research Fund of Grain Public Welfare Profession of China (Project No. 201513006), and the Major Program of Application Technology Research and Development Program of Heilongjiang Province of China (Grant No. GA15B203).

Supporting information

Supplementary Table 1—List of primers used in this study.

Supplementary Fig. 1—Product of PCR amplification of aglB gene and aglB2 gene fragment from Aspergillus niger CICC2462.

Supplementary Fig. 2—Analysis of transformants of Aspergillus niger by agarose gel electrophoresis using PCR products digested by the restriction enzyme HindIII.

Supplementary Fig. 3—Predicted N-glycosylation sites in AglB.

Supplementary Fig. 4—Minimum free energy plain structure drawing of the mRNAs encoding.

Supplementary material

10529_2018_2540_MOESM1_ESM.docx (636 kb)
Supplementary material 1 (DOCX 635 kb)

References

  1. Ahmad M, Hirz M, Pichler H, Schwab H (2014) Protein expression in Pichia pastoris: recent achievements and perspectives for heterologous protein production. Appl Microbiol Biotechnol 98(12):5301–5317CrossRefPubMedPubMedCentralGoogle Scholar
  2. Aleksieva P, Tchorbanov B, Nacheva L (2010) High-yield production of alpha-galactosidase excreted from Penicillium chrysogenum and Aspergillus niger. Biotechnol Biotechnol Equip 24(1):1620–1623CrossRefGoogle Scholar
  3. Chen Z, Yan Q, Jiang Z, Liu Y, Li Y (2015) High-level expression of a novel α-galactosidase gene from Rhizomucor miehei in Pichia pastoris and characterization of the recombinant enyzme. Protein Expr Purif 110:107–114CrossRefPubMedGoogle Scholar
  4. de Vries RP, van den Broeck HC, Dekkers E, Manzanares P, de Graaff LH, Visser J (1999) Differential expression of three α-galactosidase genes and a single β-galactosidase gene from Aspergillus niger. Appl Environ Microbiol 65(6):2453–2460PubMedPubMedCentralGoogle Scholar
  5. Gürkök S, Cekmecelioglu D, Ögel ZB (2011) Optimization of culture conditions for Aspergillus sojae expressing an Aspergillus fumigatus α-galactosidase. Bioresour Technol 102(7):4925–4929CrossRefPubMedGoogle Scholar
  6. Li J, Zhang H, Zhang Y, Shuang B, Wang D, Zhao N et al (2013) Construction of food-grade xylanase engineering strain of Aspergillus niger. J Northeast Agric Univ 44(11):7–13Google Scholar
  7. Madhavan A, Sukumaran RK (2015) Signal peptides from filamentous fungi efficiently mediate the secretion of recombinant proteins in Kluyveromyces lactis. Biochem Eng J 102:31–37CrossRefGoogle Scholar
  8. Nackley AG, Shabalina SA, Tchivileva IE, Satterfield K, Korchynskyi O, Makarov SS (2006) Human catechol-O-methyltransferase haplotypes modulate protein expression by altering mRNA secondary structure. Science 314(5807):1930–1933CrossRefPubMedGoogle Scholar
  9. Naumoff DG (2004) Phylogenetic analysis of α-galactosidases of the GH27 family. Mol Biol 38(3):388–400CrossRefGoogle Scholar
  10. Nevalainen H, Peterson R (2014) Making recombinant proteins in filamentous fungi—are we expecting too much? Front Microbiol 5:75PubMedPubMedCentralGoogle Scholar
  11. Shankar SK, Mulimani VH (2007) α-Galactosidase production by Aspergillus oryzae in solid-state fermentation. Bioresour Technol 98(4):958–961CrossRefPubMedGoogle Scholar
  12. Viana PA, de Rezende ST, Marques VM, Trevizano LM, Passos FM, Oliveira MG (2006) Extracellular α-galactosidase from Debaryomyces hansenii UFV-1 and its use in the hydrolysis of raffinose oligosaccharides. J Agric Food Chem 54(6):2385–2391CrossRefPubMedGoogle Scholar
  13. Wei SZ, Zhang H, Li J (2015) Homologous expression of asparaginase gene in Aspergillus niger. J Food Sci Biotechnol 34:554–559Google Scholar
  14. Zhang YP, Gong F, Bao GQ, Gao HW, Ji SP, Tan YX (2007) B to O erythrocyte conversion by the recombinant alpha-galactosidase. Chin Med J 120(13):1145–1150PubMedGoogle Scholar
  15. Zhang H, Wang S, Xiang Zhang X, Ji W, Song F, Zhao Y, Li J (2016) The amyR-deletion strain of Aspergillus niger CICC2462 is a suitable host strain to express secreted protein with a low background. Microb Cell Fact 15(1):68CrossRefPubMedPubMedCentralGoogle Scholar
  16. Zheng X, Fang B, Han D, Yang W, Qi F, Chen H, Li S (2016) Improving the secretory expression of an α-galactosidase from Aspergillus niger in Pichia pastoris. PLoS ONE 11(8):e0161529CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  • Yue Xu
    • 1
  • Yan-hui Wang
    • 1
  • Tian-qi Liu
    • 1
  • Hui Zhang
    • 1
  • He Zhang
    • 1
  • Jie Li
    • 1
  1. 1.College of Life ScienceNortheast Agricultural UniversityHarbinChina

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