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Variation in barley (1 → 3, 1 → 4)-β-glucan endohydrolases reveals novel allozymes with increased thermostability

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Novel barley (1 → 3, 1 → 4)-β-glucan endohydrolases with increased thermostability.

Abstract

Rapid and reliable degradation of (1 → 3, 1 → 4)-β-glucan to produce low viscosity wort is an essential requirement for malting barley. The (1 → 3, 1 → 4)-β-glucan endohyrolases are responsible for the primary hydrolysis of cell wall β-glucan. The variation in β-glucanase genes HvGlb1 and HvGlb2 that encode EI and EII, respectively, were examined in elite and exotic germplasm. Six EI and 14 EII allozymes were identified, and significant variation was found in β-glucanase from Hordeum vulgare ssp. spontaneum (wild barley), the progenitor of modern cultivated barley. Allozymes were examined using prediction methods; the change in Gibbs free energy of the identified amino acid substitutions to predict changes in enzyme stability and homology modelling to examine the structure of the novel allozymes using the existing solved EII structure. Two EI and four EII allozymes in wild barley accessions were predicted to have improved barley β-glucanase thermostability. One novel EII candidate was identified in existing backcross lines with contrasting HvGlb2 alleles from wild barley and cv Flagship. The contrasting alleles in selected near isogenic lines were examined in β-glucanase thermostability analyses. The EII from wild barley exhibited a significant increase in β-glucanase thermostability conferred by the novel HvGlb2 allele. Increased β-glucanase thermostability is heritable and candidates identified in wild barley could improve malting and brewing quality in new varieties.

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References

  • Ahokas H, Naskali L (1990) Variation of a-amylase, b-amylase, b-glucanase, pullulanase, proteinase and chitinase activity in germinated samples of the wild progenitor of barley. J I Brewing 96:27–31

    Article  CAS  Google Scholar 

  • Bamforth CW (2009) Current perspective on the role of enzymes in brewing. J Cereal Sci 50:353–357

    Article  CAS  Google Scholar 

  • Barber MG, Jackson EA, Smith DB (1994) Total and individual barley (1–3), (1–4)-b-d-glucanase activities in some green and kilned malts. J I Brewing 100:91–97

    Article  CAS  Google Scholar 

  • Dehouck Y, Kwasigroch JM, Gilis D, Rooman M (2011) PoPMuSiC 2.1: a web server for protein stability changes upon mutation and sequence optimatlity. BMC Bioinf 12:151

    Article  Google Scholar 

  • Doan DNP, Fincher GB (1992) Differences in the thermostabilities of barley (1 → 3, 1 → 4)-b-glucanase are only partially determined by N-glycosylation. FEBS Lett 309:265–271

    Article  CAS  PubMed  Google Scholar 

  • Edmunds MD, Allan GR, Fincher GB, Stuart IM (1994) The quantitation of (1 → 3, 1 → 4)-b-glucanase isoenzymes during malting using an enzyme-linked immunosorbent assay. P I Brewing 79–83

  • Eglinton JK, Langridge P, Evans DE (1998) Thermostability variation in alleles of barley beta-amylase. J Cereal Sci 28:301–309

    Article  CAS  Google Scholar 

  • European Brewery Convention (EBC), 1997. Analytica – EBC. Verlag Hands Carl Getränke-Fachverlag, Nurnberg, Germany

  • Evans DE, Sheehan MC, Stewart DC (1999) The impact of malt derived proteins on beer foam quality. Part II: the influence of malt foam-positive proteins and non-starch polysaccharides on beer foam quality. J I Brewing 105:171–178

    Article  CAS  Google Scholar 

  • Georg-Kraemer JE, Caierão E, Minella E, Barbosa-Neto JF, Cavalli SS (2004) The (1–3,1–4)-β-glucanases in malting barley: enzyme survival and genetic and environmental effects. J I Brewing 110:303–308

    Article  CAS  Google Scholar 

  • Jensen LG, Olsen O, Kops O, Wolf N, Thomsen KK, von Wettstein D (1996) Transgenic barley expressing a protein-engineered, thermostable (1,3–1,4)-b-glucanase during germination. Proc Natl Acad Sci USA 93:3487–3491

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jin X, Cai S, Han Y, Wang J, Wei K, Zhang G (2011) Genetic variants of HvGlb1 in Tibetan annual wild barley and cultivated barley and their correlation with malt quality. J Cereal Sci 53:59–64

    Article  CAS  Google Scholar 

  • Karakousis A, Langridge P (2003) A high-throughput plant DNA extraction method for marker analysis. Plant Mol Biol Rep 21:95–95

    Article  Google Scholar 

  • Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, Buxton S, Cooper A, Markowitz S, Duran C, Thierer T, Ashton B, Mentjies P, Drummond A (2012) Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 28:1647–1649

    Article  PubMed  PubMed Central  Google Scholar 

  • Litts JC, Simmons CR, Karrer EE, Huang N, Rodriguez RL (1990) The isolation and characterization of a barley 1,3–1,4-b-glucanase gene. Eur J Biochem 194:831–838

    Article  CAS  PubMed  Google Scholar 

  • Loi L, Barton PA, Fincher GB (1987) Survival of barley (1→3, 1→4)-b-glucanase isoenzymes during kilning and mashing. J Cereal Sci 5:45–50

    Article  CAS  Google Scholar 

  • Lu F, Lipka AE, Glaubitz J, Elshire R, Cherney JH, Casler MD, Buckler ES, Costich DE (2013) Switchgrass genomic diversity, ploidy, and evolution: novel insights from a network-based SNP discovery protocol. PLoS Genet 9(1):e1003215

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • MacLeod LC, Lance RCM, Brown AHD (1991) Chromosomal mapping of the Glb 1 locus encoding (1→3,1→4)-b-d-glucan 4-glucanohydrolase EI in barley. J Cereal Sci 13:291–298

    Article  CAS  Google Scholar 

  • Matthews BW, Nicholson H, Becktel WJ (1987) Enhanced protein thermostability from site-directed mutations that decrease the entropy of unfolding. Proc Natl Acad Sci USA 84:6663–6667

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • McCleary BV, Shameer I (1987) Assay of malt b-glucanase using azo-barley glucan—an improved precipitant. J I Brewing 93:87–90

    Article  CAS  Google Scholar 

  • Müller JJ, Thomsen KK, Heinemann U (1998) Crystal Structure of barley 1,3–1,4-b-glucanase at 2.0-Å resolution and comparison with Bacillus 1,3–1,4-b-glucanase. J Biol Chem 273:3438–3446

    Article  PubMed  Google Scholar 

  • Nevo E, Zohary D, Brown AHD, Haber M (1979) Genetic diversity and environmental association of wild barley, Hordeum spontaneum, in Israel. Evol Int J org Evol 33:815–833

    Article  CAS  Google Scholar 

  • Poland JA, Brown PJ, Sorrells ME, Jannink J-L (2012) Development of high-density genetic maps for barley and wheat using a novel two-enzyme genotyping-by-sequencing approach. PLOS One 7(2): e32253

  • Querol E, Perez-Pons JA, Mozo-Villarias A (1996) Analysis of protein conformational characteristics related to thermostability. Protein Eng 9:265–271

    Article  CAS  PubMed  Google Scholar 

  • Sali A, Blundell TL (1993) Comparative protein modelling by satisfaction of spatial restraints. J Mol Biol 234:779–815

    Article  CAS  PubMed  Google Scholar 

  • Slakeski N, Fincher GB (1992) Developmental Regulation of (1 → 3, 1 → 4)-b-glucanase gene expression in barley—tissue-specific expression of individual isoenzymes. Plant Physiol 99:1226–1231

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Slakeski N, Baulcombe DC, Devos KM, Ahluwalia B, Doan DNP, Fincher GB (1990) Structure and tissue-specific regulation of genes encoding barley (1 → 3, 1 → 4)-b-glucan endohydrolases. Mol Gen Genet 224:437–449

    Article  CAS  PubMed  Google Scholar 

  • Stewart RJ, Varghese JN, Garrett TPJ, Høj PB, Fincher GB (2001) Mutant barley (1 → 3, 1 → 4)-b-glucan endohydrolases with enhanced thermostability. Protein Eng 14:245–253

    Article  CAS  PubMed  Google Scholar 

  • Stuart IM, Loi L, Fincher GB (1988) Varietal and environmental variations in (1 → 3, 1 → 4)-b-glucan levels and (1 → 3, 1 → 4)-b-glucanase potential in barley: relationships to malting quality. J Cereal Sci 7:61–71

    Article  CAS  Google Scholar 

  • Teng D, Wang J-h, Fan Y, Yang Y-l, Tian Z-g, Luo J, Yang G-p, Zhang F (2006) Cloning of b-1,3–1,4-glucanase gene from Bacillus licheniformis EGW039 (CGMCC 0635) and its expression in Escherichia coli BL21 (DE3). Appl Microbiol Biotechnol 72:705–712

    Article  CAS  PubMed  Google Scholar 

  • Tsai L-C, Chen Y-N, Shyur L-F (2008) Structural modeling of glucanase–substrate complexes suggests a conserved tyrosine is involved in carbohydrate recognition in plant 1,3–1,4-b-d-glucanases. J Comput Aided Mol Des 22:915–923

    Article  CAS  PubMed  Google Scholar 

  • Wolf N (1991) Complete Nucleotide Sequence of a Hordeum vulgare gene encoding (1 → 3, 1 → 4)-b-glucanase isoenzyme II. Plant Physiol 96:1382–1384

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wolf N (1992) Structure of the genes encoding Hordeum vulgare (1→3,1→4)-b-glucanase isoenzymes I and II and functional analysis of their promoters in barley aleurone protoplasts. Mol Gen Genet 234:33–42

    CAS  PubMed  Google Scholar 

  • Woodward JR, Fincher GB (1982) Substrate specificities and kinetic-properties of two (1→3,1→4)-b-d.-glucan endo-hydrolases from germinating barley (Hordeum vulgare). Carbohyd Res 106:111–122

  • Zwickert-Menteur S, Jestin L, Branlard G (1996) Amy2 polymorphism as a possible marker of β-glucanase activity in barley (Hordeum vulgare L). J Cereal Sci 24:55–63

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Acknowledgements

This research received financial support from Australian farmers and the Federal Government through the Grains Research and Development Corporation (GRS10407), the Australian Research Council Centre of Excellence in Plant Cell Walls (CE110001007) and an Australian Postgraduate Award through the University of Adelaide.

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Authors and Affiliations

  1. School of Agriculture, Food and Wine, The University of Adelaide, Waite Campus, PMB 1, Glen Osmond, SA, 5064, Australia

    Juanita C. Lauer & Jason K. Eglinton

  2. Australian Research Council Centre of Excellence in Plant Cell Walls, School of Agriculture, Food and Wine, The University of Adelaide, Waite Campus, PMB 1, Glen Osmond, SA, 5064, Australia

    Suong Cu & Rachel A. Burton

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  1. Juanita C. Lauer
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  2. Suong Cu
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  3. Rachel A. Burton
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  4. Jason K. Eglinton
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Correspondence to Juanita C. Lauer.

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Communicated by Gary Muehlbauer.

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Lauer, J.C., Cu, S., Burton, R.A. et al. Variation in barley (1 → 3, 1 → 4)-β-glucan endohydrolases reveals novel allozymes with increased thermostability. Theor Appl Genet 130, 1053–1063 (2017). https://doi.org/10.1007/s00122-017-2870-z

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  • DOI: https://doi.org/10.1007/s00122-017-2870-z

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