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Applied Microbiology and Biotechnology

, Volume 102, Issue 23, pp 10027–10041 | Cite as

Secretory expression of β-mannanase in Saccharomyces cerevisiae and its high efficiency for hydrolysis of mannans to mannooligosaccharides

  • Junquan Liu
  • Abdul Basit
  • Ting Miao
  • Fengzhen Zheng
  • Hang Yu
  • Yan Wang
  • Wei JiangEmail author
  • Yunhe CaoEmail author
Biotechnologically relevant enzymes and proteins

Abstract

Degradation of mannans is a key process in the production of foods and prebiotics. β-Mannanase is the key enzyme that hydrolyzes 1,4-β-D-mannosidic linkages in mannans. Heterogeneous expression of β-mannanase in Pichia pastoris systems is widely used; however, Saccharomyces cerevisiae expression systems are more reliable and safer. We optimized β-mannanase gene from Aspergillus sulphureus and expressed it in five S. cerevisiae strains. Haploid and diploid strains, and strains with constitutive promoter TEF1 or inducible promoter GAL1, were tested for enzyme expression in synthetic auxotrophic or complex medium. Highest efficiency expression was observed for haploid strain BY4741 integrated with β-mannanase gene under constitutive promoter TEF1, cultured in complex medium. In fed-batch culture in a fermentor, enzyme activity reached ~ 24 U/mL after 36 h, and production efficiency reached 16 U/mL/day. Optimal enzyme pH was 2.0–7.0, and optimal temperature was 60 °C. In studies of β-mannanase kinetic parameters for two substrates, locust bean gum galactomannan (LBG) gave Km = 24.13 mg/mL and Vmax = 715 U/mg, while konjac glucomannan (KGM) gave Km = 33 mg/mL and Vmax = 625 U/mg. One-hour hydrolysis efficiency values were 57% for 1% LBG, 74% for 1% KGM, 39% for 10% LBG, and 53% for 10% KGM. HPLC analysis revealed that the major hydrolysis products were the oligosaccharides mannose, mannobiose, mannotriose, mannotetraose, mannopentaose, and mannohexaose. Our findings show that this β-mannanase has high efficiency for hydrolysis of mannans to mannooligosaccharides, a type of prebiotic, suggesting strong potential application in food industries.

Keywords

β-Mannanase Expression Saccharomyces cerevisiae Mannans Mannooligosaccharides 

Notes

Acknowledgments

This study was supported by Scientific and Technical Supporting Programmes (2013BAD10B01). The authors are grateful to Dr. S. Anderson for the English editing of the manuscript and Prof. Huiqiang Lou for donating strains.

Compliance with ethical standards

Conflict of interest

The authors declare that have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants performed by any of the authors.

Supplementary material

253_2018_9355_MOESM1_ESM.pdf (424 kb)
ESM 1 (PDF 424 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Beijing Advanced Innovation Center for Food Nutrition and Human Health, State Key Laboratory of Agro-BiotechnologyChina Agricultural UniversityBeijingChina
  2. 2.Liaoning Union Pharmaceutical Company LimitedBenxiChina
  3. 3.State Key Laboratory of Animal NutritionChina Agricultural UniversityBeijingChina

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