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Efficient pullulan production by bioconversion using Aureobasidium pullulans as the whole-cell catalyst

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Abstract

In this study, pullulan production was achieved by whole-cell bioconversion with Aureobasidium pullulans CCTCC M 2012259. Response surface methodology was applied to optimize the seed medium for incubating cells with high capability of pullulan bioconversion. Three medium components, namely, yeast extract, MgSO4·7H2O, and glucose were identified by Plackett-Berman design as significant factors affecting the cells’ pullulan bioconversion capability. A three-level Box-Behnken design was then employed to determine the optimal levels of the three components. A mathematical model was developed to show the influence of each medium component and its effects on the cells’ pullulan bioconversion capability. The model predicted a maximum pullulan bioconversion capability of 32.28 mg/g/h at the optimal yeast extract, MgSO4·7H2O, and glucose concentrations of 3.57, 0.18, and 63.97 g/l, respectively. The validation experiments showed that the cells’ pullulan bioconversion capability was improved by 23.1 % when the optimal medium was used, as compared with that obtained with the basic medium. Subsequently, the gene expression and activities of the key enzymes involved in pullulan biosynthesis were evaluated. When the optimal medium was employed, the transcriptional levels of pgm1 and fks were up-regulated by 2.5- and 1.2-fold, respectively, and the α-phosphoglucose mutase and glucosyltransferase activities were increased by 17 and 19 %, respectively, when compared with those achieved using the basic medium. These results indicated that pullulan bioconversion using A. pullulans CCTCC M 2012259 as the whole-cell catalyst is an attractive approach for efficient pullulan production and can be applied for the production of other polysaccharides.

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References

  • Box GEP, Behnken DW (1960) Some new three level designs for the study of quantitative variables. Technometrics 2:455–475

    Article  Google Scholar 

  • Cheng KC, Demirci A, Catchmark JM (2010) Effects of plastic composite support and pH profiles on pullulan production. Appl Microbiol Biotechnol 86:853–861

    Article  CAS  PubMed  Google Scholar 

  • Cheng KC, Demirci A, Catchmark JM (2011a) Pullulan: biosynthesis, production, and applications. Appl Microbiol Biotechnol 92:29–44

    Article  CAS  PubMed  Google Scholar 

  • Cheng KC, Demirci A, Catchmark MJ (2011b) Continuous pullulan fermentation in biofilm reactor by Aureobasidium pullulans. Appl Microbiol Biotechnol 90:921–927

    Article  CAS  PubMed  Google Scholar 

  • Chi Z, Wang F, Chi Z, Yue L, Liu G, Zhang T (2009) Bioproducts from Aureobasidium pullulans, a biotechnologically important yeast. Appl Microbiol Biotechnol 82:793–804

    Article  CAS  PubMed  Google Scholar 

  • Choudhury AR, Bhattacharyya MS, Prasad GS (2012) Application of response surface methodology to understand the interaction of media components during pullulan production by Aureobasidium pullulans RBF-4A3. Biocatal Agric Biotechnol 1:232–237

    CAS  Google Scholar 

  • D’Haene B, Vandesompele J, Hellemans J (2010) Accurate and objective copy number profiling using real-time quantitative PCR. Methods 50:262–270

    Article  PubMed  Google Scholar 

  • de Carvalho CC (2011) Enzymatic and whole cell catalysis: finding new strategies for old processes. Biotechnol Adv 29:75–83

    Article  PubMed  Google Scholar 

  • Duan X, Chi Z, Wang L, Wang X (2008) Influence of different sugars on pullulan production and activities of α-phosphoglucose mutase, UDPG-pyrophosphorylase and glucosyltransferase involved in pullulan synthesis in Aureobasidium pullulans Y68. Carbohyd Polym 73:587–593

    Article  CAS  Google Scholar 

  • Farris S, Unalan IU, Introzzi L, Fuentes-Alventosa JM, Cozzolino CA (2014) Pullulan-based films and coatings for food packaging: present applications, emerging opportunities, and future challenges. J Appl Polym Sci. doi:10.1002/APP.40539

    Google Scholar 

  • Gaur R, Singh R, Gupta M, Gaur MK (2010) Aureobasidium pullulans, an economically important polymorphic yeast with special reference to pullulan. Afr J Biotechnol 9:7989–7997

    CAS  Google Scholar 

  • Guo M, Hu CH (2010) Bioconversion: from whole cell biocatalysis to metabolic engineering. China Biotechnol 30(4):110–115

    CAS  Google Scholar 

  • Jiang L (2010) Optimization of fermentation conditions for pullulan production by Aureobasidium pullulans using response surface methodology. Carbohyd Polym 79:414–417

    Article  CAS  Google Scholar 

  • Jiang LF, Wu SJ, Kim JM (2011) Effect of different nitrogen sources on activities of UDPG-pyrophosphorylase involved in pullulan synthesis and pullulan production by Aureobasidium pullulans. Carbohyd Polym 86:1085–1088

    Article  CAS  Google Scholar 

  • Kang JX, Chen XJ, Chen WR, Li MS, Fang Y, Li DS, Ren YZ, Liu DQ (2011) Enhanced production of pullulan in Aureobasidium pullulans by a new process of genome shuffling. Process Biochem 46:792–795

    Article  CAS  Google Scholar 

  • Kang TS, Korber DR, Tanaka T (2013) Contributions of citrate in redox potential maintenance and ATP production: metabolic pathways and their regulation in Lactobacillus panis PM1. Appl Microbiol Biotechnol 97:8693–8703

    Article  CAS  PubMed  Google Scholar 

  • Khuri AI, Mukhopadhyay S (2010) Response surface methodology. WIREs Comput Stat 2:128–149

    Article  Google Scholar 

  • Leathers TD (2003) Biotechnological production and applications of pullulan. Appl Microbiol Biotechnol 62:468–473

    Article  CAS  PubMed  Google Scholar 

  • Liu GQ, Wang XL (2007) Optimization of critical medium components using response surface methodology for biomass and extracellular polysaccharide production by Agaricus blazei. Appl Microbiol Biotechnol 74:78–83

    Article  CAS  PubMed  Google Scholar 

  • Ma ZC, Fu WJ, Liu GL, Wang ZP, Chi ZM (2014) High-level pullulan production by Aureobasidium pullulans var. melanogenium P16 isolated from mangrove system. Appl Microbiol Biotechnol 98:4865–4873

    Article  CAS  PubMed  Google Scholar 

  • Mulchandani A, Luong JHT, LeDuy A (1989) Biosynthesis of pullulan using immobilized Aureobasidium pullulans cells. Biotechnol Bioeng 33:306–312

    Article  CAS  PubMed  Google Scholar 

  • Plackett RL, Burman JP (1946) The design of optimum multi-factorial experiments. Biometrika 33:305–325

    Article  Google Scholar 

  • Pscheidt B, Glieder A (2008) Yeast cell factories for fine chemical and API production. Microb Cell Fact 7:25

    Article  PubMed Central  PubMed  Google Scholar 

  • Schoemaker HE, Mink D, Wubbolts MG (2003) Dispelling the myths-biocatalysis in industrial synthesis. Science 299:1694–1697

    Article  CAS  PubMed  Google Scholar 

  • Seo HP, Son CW, Chung CH, Jung DI, Kim SK, Gross RA, Kaplan DL, Lee JW (2004) Production of high molecular weight pullulan by Aureobasidium pullulans HP-2001 with soybean pomace as a nitrogen source. Bioresource Tech 95:293–299

    Article  CAS  Google Scholar 

  • Seo HP, Jo KI, Son CW, Yang JK, Chung CH, Nam SW, Kim SK, Lee JW (2006) Continuous production of pullulan by Aureobasidium pullulans HP-2001 with feeding of high concentration of sucrose. J Microbiol Biotechnol 16:374–380

    CAS  Google Scholar 

  • Singh RS, Saini GK, Kennedy JF (2008) Pullulan: microbial sources, production and applications. Carbohyd Polym 73:515–531

    Article  CAS  Google Scholar 

  • Singh R, Gaur R, Tiwari S, Gaur MK (2012) Production of pullulan by a thermotolerant Aureobasidium pullulans strain in nonstirred fed batch fermentation process. Braz J Microbiol 43:1042–1050

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Wang D, Yu X, Wei G (2013) Pullulan production and physiological characteristics of Aureobasidium pullulans under acid stress. Appl Microbiol Biotechnol 97:8069–8077

    Article  CAS  PubMed  Google Scholar 

  • Wang D, Ju X, Zhou D, Wei G (2014) Efficient production of pullulan using rice hull hydrolysate by adaptive laboratory evolution of Aureobasidium pullulans. Bioresource Tech 164:12–19

    Article  CAS  Google Scholar 

  • West TP (2000) Exopolysaccharide production by entrapped cells of the fungus Aureobasidium pullulans ATCC 201253. J Basic Microbiol 40:397–401

    Article  CAS  PubMed  Google Scholar 

  • West TP (2010) Pullulan production by Aureobasidium pullulans cells immobilized on ECTEOLA-cellulose. Ann Microbiol 60:763–766

    Article  CAS  Google Scholar 

  • Wu S, Chen J, Pan S (2012) Optimization of fermentation conditions for the production of pullulan by a new strain of Aureobasidium pullulans isolated from sea mud and its characterization. Carbohyd Polym 87:1696–1700

    Article  CAS  Google Scholar 

  • Yang CG, Wang XL, Tian J, Liu W, Wu F, Jiang M, Wen H (2013) Evaluation of reference genes for quantitative real-time RT-PCR analysis of gene expression in Nile tilapia (Oreochromis niloticus). Gene 527:183–192

    Article  CAS  PubMed  Google Scholar 

  • Yoshida H, Hara KY, Kiriyama K, Nakayama H, Okazaki F, Matsuda F, Ogino C, Fukuda H, Kondo A (2011) Enzymatic glutathione production using metabolically engineered Saccharomyces cerevisiae as a whole-cell biocatalyst. Appl Microbiol Biotechnol 91:1001–1006

    Article  CAS  PubMed  Google Scholar 

  • Youssef F, Roukas T, Biliaderis CG (1999) Pullulan production by a non-pigmented strain of Aureobasidium pullulans using batch and fed-batch culture. Process Biochem 34:355–366

    Article  CAS  Google Scholar 

  • Yu X, Wang Y, Wei G, Dong Y (2012) Media optimization for elevated molecular weight and mass production of pigment-free pullulan. Carbohyd Polym 89:928–934

    Article  CAS  Google Scholar 

  • Zhang HT, Zhan XB, Zheng ZY, Wu JR, Yu XB, Lin CC, Jiang Y (2011) Sequence and transcriptional analysis of the genes responsible for curdlan biosynthesis in Agrobacterium sp. ATCC 31749 under simulated dissolved oxygen gradients conditions. Appl Microbiol Biotechnol 91:163–175

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was financially supported by the National Natural Science Foundation of China (21376155). D. Cao was subsidized by Hui-Chun Chin and Tsung-Dao Lee Chinese Undergraduate Research Endowment. G. Wei was sponsored by Qing Lan Project of Jiangsu Province.

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Correspondence to Gong-Yuan Wei.

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Da-Hui Wang and Xiao-Min Ju contributed equally to this work.

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Ju, XM., Wang, DH., Zhang, GC. et al. Efficient pullulan production by bioconversion using Aureobasidium pullulans as the whole-cell catalyst. Appl Microbiol Biotechnol 99, 211–220 (2015). https://doi.org/10.1007/s00253-014-6100-1

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