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Cloning and characterization of a novel fructan 6-exohydrolase strongly inhibited by sucrose in Lolium perenne

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Abstract

Main conclusion

The first 6-fructan exohydrolase (6-FEH) cDNA from Lolium perenne was cloned and characterized. Following defoliation, Lp6 - FEHa transcript level unexpectedly decreased together with an increase in total FEH activity.

Lolium perenne is a major forage grass species that accumulates fructans, mainly composed of β(2,6)-linked fructose units. Fructans are mobilized through strongly increased activities of fructan exohydrolases (FEHs), sustaining regrowth following defoliation. To understand the complex regulation of fructan breakdown in defoliated grassland species, the objective was to clone and characterize new FEH genes in L. perenne. To find FEH genes related to refoliation, a defoliated tiller base cDNA library was screened. Characterization of the recombinant protein was performed in Pichia pastoris. In this report, the cloning and enzymatic characterization of the first 6-FEH from L. perenne is described. Following defoliation, during fructan breakdown, Lp6-FEHa transcript level unexpectedly decreased in elongating leaf bases (ELB) and in mature leaf sheaths (tiller base) in parallel to increased total FEH activities. In comparison, transcript levels of genes coding for fructosyltransferases (FTs) involved in fructan biosynthesis also decreased after defoliation but much faster than FEH transcript levels. Since Lp6-FEHa was strongly inhibited by sucrose, mechanisms modulating FEH activities are discussed. It is proposed that differences in the regulation of FEH activity among forage grasses influence their tolerance to defoliation.

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Abbreviations

6G-FFT:

6G-Fructan:fructan fructoyltransferase

6-SFT:

6-Sucrose:fructan fructosyltransferase

DP:

Degree of polymerization

ELB:

Elongating leaf bases

ES:

External sheath

FEH:

Fructan exohydrolase

FT:

Fructosyltransferase

IS:

Internal sheath

MS:

Middle sheath

SST:

Sucrose:sucrose fructosyltransferase

WSC:

Water-soluble carbohydrates

References

  • Asega AF, do Nascimento JR, Schroeven L, Van Den Ende W, Carvalho MA (2008) Cloning, characterization and functional analysis of a 1-FEH cDNA from Vernonia herbacea (Vell.) Rusby. Plant Cell Physiol 49:1185–1195

    CAS  PubMed  Google Scholar 

  • Asega AF, do Nascimento JR, Carvalho MA (2011) Increased expression of fructan 1-exohydrolase in rhizophores of Vernonia herbacea during sprouting and exposure to low temperature. J Plant Physiol 168:558–565

    Article  CAS  PubMed  Google Scholar 

  • Bachmann M, Matile P, Keller F (1994) Metabolism of the raffinose family oligosaccharides in leaves of Ajuga reptans L. Plant Physiol 105:1335–1345

    CAS  PubMed Central  PubMed  Google Scholar 

  • Ballard RA, Simpson RJ, Pearce GR (1990) Losses of the digestible components of annual ryegrass (Lolium rigidum Gaudin) during senescence. Aust J Agr Res 41:719–731

    Article  CAS  Google Scholar 

  • Bendtsen JD, Nielsen H, von Heijne G, Brunak S (2004) Improved prediction of signal peptides: signalP 3.0. J Mol Biol 340:783–795

    Article  PubMed  Google Scholar 

  • Blake JD, Clarke ML, Jansson PE, McNeil KE (1982) Fructan from Erwinia herbicola. J Bacteriol 151:1595–1597

    CAS  PubMed Central  PubMed  Google Scholar 

  • Cairns AJ, Ashton JE (1993) Species-dependent patterns of fructan synthesis by enzymes from excised leaves of oat, wheat, barley and timothy. New Phytol 124:381–388

    Article  CAS  Google Scholar 

  • Casler MD (2005) Agronomic performance of timothy germplasm from forage and turf sods under two harvest managements. Crop Sci 45:1990–1996

    Article  Google Scholar 

  • Chatterton NJ, Harrison PA, Thornley WR, Draper EA (1990) Oligosaccharides in foliage of Agropyron, Bromus, Dactylis, Festuca, Lolium and Phleum. New Phytol 114:167–171

    Article  CAS  Google Scholar 

  • Coller J, Parker R (2004) Eukaryotic mRNA decapping. Annu Rev Biochem 73:861–890

    Article  CAS  PubMed  Google Scholar 

  • De Coninck B, Le Roy K, Francis I, Clerens S, Vergauwen R, Halliday AM, Smith SM, Van Laere A, Van den Ende W (2005) Arabidopsis AtcwINV3 and 6 are not invertases but are fructan exohydrolases (FEHs) with different substrate specificities. Plant Cell Environ 28:432–443

    Article  Google Scholar 

  • De Roover J, De Winter M, Van Laere A, Timmermans J, Van den Ende W (1999) Purification and properties of a second fructan exohydrolase from the roots of Cichorium intybus L. Physiol Plant 106:28–34

    Article  Google Scholar 

  • Del Viso F, Puebla AF, Hopp HE, Heinz RA (2009) Cloning and functional characterization of a fructan 1-exohydrolase (1-FEH) in the cold tolerant patagonian species Bromus pictus. Planta 231:13–25

    Article  PubMed  Google Scholar 

  • Hendry G (1993) Evolutionary origins and natural functions of fructans: a climatological, biogeographic and mechanistic appraisal. New Phytol 123:3–14

    Article  CAS  Google Scholar 

  • Henson CA, Livingston DP III (1996) Purification and characterization of an oat fructan exohydrolase that preferentially hydrolyzes β-2,6-fructans. Plant Physiol 110:639–644

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Henson CA, Livingston DP III (1998) Characterization of a fructan exohydrolase purified from barley stems that hydrolyzes multiple fructofuranosidic linkages. Plant Physiol Biochem 36:715–720

    CAS  Google Scholar 

  • Hincha DK, Zuther E, Hellwege EM, Heyer AG (2002) Specific effects of fructo- and gluco-oligosaccharides in the preservation of liposomes during drying. Glycobiology 12:103–110

    Article  CAS  PubMed  Google Scholar 

  • Joudi M, Ahmadi A, Mohamadi V, Abbasi A, Vergauwen R, Mohamadi H, Van den Ende W (2012) Comparison of fructan dynamics in two wheat cultivars with different capacities of accumulation and remobilization under terminal drought stress. Physiol Plant 144:1–12

    Article  CAS  PubMed  Google Scholar 

  • Kawakami A, Yoshida M, Van den Ende W (2005) Molecular cloning and functional analysis of a novel 6&1-FEH from wheat (Triticum aestivum L.) preferentially degrading small graminans like bifurcose. Gene 358:93–101

    Article  CAS  PubMed  Google Scholar 

  • Kawakami A, Sato Y, Yoshida M (2008) Genetic engineering of rice capable of synthesizing fructans and enhancing chilling tolerance. J Exp Bot 59:793–802

    Article  CAS  PubMed  Google Scholar 

  • Kusch U, Harms K, Rausch T, Greiner S (2009) Inhibitors of plant invertases do not affect the structurally related enzymes of fructan metabolism. New Phytol 181:601–612

    Article  CAS  PubMed  Google Scholar 

  • Lammens W, Le Roy K, Schroeven L, Van Laere A, Rabjins A, Van den Ende W (2009) Structural insights into glycoside hydrolase family 32 and 68 enzymes: functional implications. J Exp Bot 60:727–740

    Article  CAS  PubMed  Google Scholar 

  • Lasseur B, Lothier J, Djoumad A, De Coninck B, Smeekens S, Van Laere A, Morvan-Bertrand A, Van den Ende W, Prud’homme M-P (2006) Molecular and functional characterization of a cDNA encoding fructan:fructan 6G-fructosyltransferase (6G-FFT)/fructan:fructan 1-fructosyltransferase (1-FFT) from perennial ryegrass (Lolium perenne L). J Exp Bot 57:2719–2734

    Article  CAS  PubMed  Google Scholar 

  • Lasseur B, Lothier J, Wiemken A, Van Laere A, Morvan-Bertrand A, Van den Ende W, Prud’homme M-P (2011) Towards a better understanding of the generation of fructan structure diversity in plants: molecular and functional characterization of a sucrose:fructan 6-fructosyltransferase (6-SFT) cDNA from perennial ryegrass (Lolium perenne). J Exp Bot 62:1871–1885

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lavut A, Raveh D (2012) Sequestration of highly expressed mRNAs in cytoplasmic granules, P-bodies, and stress granules enhances cell viability. PLoS Genet 8:e1002527

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Le Roy K, Lammens W, Verhaest M, De Coninck B, Rabijns A, Van Laere A, Van den Ende W (2007) Unraveling the difference between invertases and fructan exohydrolases: a single amino acid (Asp-239) substitution transforms Arabidopsis cell wall invertase1 into a fructan 1-exohydrolase. Plant Physiol 145:616–625

    Article  PubMed Central  PubMed  Google Scholar 

  • Le Roy K, Lammens W, Van Laere A, Van den Ende W (2008) Influencing the binding configuration of sucrose in the active site of chicory fructan 1-exohydrolase and sugar beet fructan 6-exohydrolase. New Phytol 178:572–580

    Article  PubMed  Google Scholar 

  • Le Roy K, Vergauwen R, Struyf T, Yuan S, Lammens W, Matrai J, De Maeyer M, Van den Ende W (2013) Understanding the role of defective invertase in plants: tobacco Nin88 fails to degrade sucrose. Plant Physiol 161:1670–1681

    Article  PubMed Central  PubMed  Google Scholar 

  • Lee JM, Donaghy DJ, Roche JR (2008) Effect of defoliation severity on regrowth and nutritive value of perennial ryegrass dominant swards. Agron J 100:308–314

    Article  Google Scholar 

  • Livingston DP III, Henson CA (1998) Apoplastic sugars, fructans, fructan exohydrolase, and invertase in winter oat: responses to second-phase cold hardening. Plant Physiol 116:403–408

    Article  CAS  PubMed Central  Google Scholar 

  • Livingston DP III, Hincha DK, Heyer AG (2009) Fructan and its relationship to abiotic stress tolerance in plants. Cell Mol Life Sci 66:2007–2023

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lothier J, Lasseur B, Djoumad A, Le Roy K, Van Laere A, Prud’homme M-P, Barre P, Van den Ende W, Morvan-Bertrand A (2007) Cloning, gene mapping and functional analysis of a fructan 1-exohydrolase (1-FEH) from Lolium perenne rather implicated in fructan synthesis than in fructan mobilization. J Exp Bot 58:1969–1983

    Article  CAS  PubMed  Google Scholar 

  • Lothier J, Lasseur B, Morvan-Bertrand A, Prud’homme M-P (2010) Hexokinase dependent sugar signaling represses fructan exohydrolase (FEH) activity in Lolium perenne. Funct Plant Biol 37:1151–1160

    Article  CAS  Google Scholar 

  • Marx SP, Nösberger J, Frehner M (1997) Hydrolysis of fructan in grasses: a β-(2-6)-linkage specific fructan-beta-fructosidase from stubble of Lolium perenne. New Phytol 135:279–290

    Article  CAS  Google Scholar 

  • Michiels A, Van Laere A, Van den Ende W, Tucker M (2004) Expression analysis of a chicory fructan 1-exohydrolase gene reveals complex regulation by cold. J Exp Bot 55:1325–1333

    Article  CAS  PubMed  Google Scholar 

  • Morvan A, Challe G, Prud’homme M-P, Le Saos J, Boucaud J (1997) Rise of fructan exohydrolase activity in stubble of Lolium perenne after defoliation is decreased by uniconazole, an inhibitor of the biosynthesis of gibberellins. New Phytol 136:81–88

    Article  CAS  Google Scholar 

  • Morvan-Bertrand A, Boucaud J, Prud’homme M-P (1999) Influence of initial levels of carbohydrates, fructans, nitrogen, and soluble proteins on regrowth of Lolium perenne L. cv. bravo following defoliation. J Exp Bot 50:1817–1826

    Article  CAS  Google Scholar 

  • Morvan-Bertrand A, Boucaud J, Le Saos J, Prud’homme M-P (2001) Roles of the fructans from leaf sheaths and from the elongating leaf bases in the regrowth following defoliation of Lolium perenne L. Planta 213:109–120

    Article  CAS  PubMed  Google Scholar 

  • Nagaraj VJ, Altenbach D, Galati V, Lüscher M, Meyer AD, Boller T, Wiemken A (2004) Distinct regulation of sucrose:sucrose-1-fructosyltransferase (1-SST) and sucrose:fructan-6-fructosyltransferase (6-SFT), the key enzymes of fructan synthesis in barley leaves: 1-SST as the pacemaker. New Phytol 161:735–748

    Article  CAS  Google Scholar 

  • Pavis N, Chatterton NJ, Harrison PA, Baumgartner S, Praznik W, Boucaud J, Prud’homme M-P (2001) Structure of fructans in roots and leaf tissues of Lolium perenne. New Phytol 150:83–95

    Article  CAS  Google Scholar 

  • Peshev D, Van den Ende W (2014) Fructans: prebiotics and immunomodulators. J Funct Foods 8:348–357

    Article  CAS  Google Scholar 

  • Peshev D, Vergauwen R, Moglia A, Hideg E, Van den Ende W (2013) Towards understanding vacuolar antioxidant mechanisms: a role for fructans? J Exp Bot 64:1025–1038

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Pilon-Smits EAH, Terry N, Sears T, van Dun K (1999) Enhanced drought resistance in fructan-producing sugar beet. Plant Physiol Biochem 37:313–317

    CAS  Google Scholar 

  • Pollock CJ (1986) Fructans and the metabolism of sucrose in vascular plants. New Phytol 104:1–24

    Article  CAS  Google Scholar 

  • Prud’homme M-P, Gonzalez B, Billard J, Boucaud J (1992) Carbohydrate content, fructan and sucrose enzyme activities in roots, stubble and leaves of ryegrass (Lolium perenne L.) as affected by source/sink modification after cutting. J Plant Physiol 140:282–291

    Article  Google Scholar 

  • Rausch T, Greiner S (2004) Plant protein inhibitors of invertases. Bioch Biophys Acta 1696:253–261

    CAS  Google Scholar 

  • Schnyder H (1993) The role of carbohydrate storage and redistribution in the source-sink relations of wheat and barley during grain filling: a review. New Phytol 123:233–245

    Article  Google Scholar 

  • Shiomi N (1981) Purification and characterisation of 6G-fructosyltransferase from the roots of asparagus (Asparagus officinalis L.). Carbohydr Res 96:281–292

    Article  CAS  Google Scholar 

  • Smeekens S, Ma J, Hanson J, Rolland F (2010) Sugar signals and molecular networks controlling plant growth. Curr Opin Plant Biol 13:274–279

    Article  CAS  PubMed  Google Scholar 

  • Sprenger N, Bortlik K, Brandt A, Boller T, Wiemken A (1995) Purification, cloning, and functional expression of sucrose:fructan 6-fructosyltransferase, a key enzyme of fructan synthesis in barley. Proc Natl Acad Sci USA 92:11652–11656

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Tamura KI, Sanada Y, Tase K, Komatsu T, Yoshida M (2011) Pp6-FEH1 encodes an enzyme for degradation of highly polymerized levan and is transcriptionally induced by defoliation in timothy (Phleum pratense L.). J Exp Bot 62:3421–3431

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Van den Ende W (2013) Multifunctional fructans and raffinose family oligosaccharides. Front Plant Sci 4:1–10

    Article  Google Scholar 

  • Van den Ende W, El-Esawe SK (2013) Sucrose signalling pathways leading to fructan and anthocyanin accumulation: a dual function in abiotic and biotic stress responses? Environ Exp Bot. doi:10.1016/j.envexpbot.2013.09.017 (In Press)

    Google Scholar 

  • Van den Ende W, Van Laere A (1996) Fructan synthesizing and degrading activities in chicory roots (Cichorium intybus L) during growth, storage and forcing. J Plant Physiol 149:43–50

    Article  Google Scholar 

  • Van den Ende W, Michiels A, De Roover J, Verhaert P, Van Laere A (2000) Cloning and functional analysis of chicory root fructan 1-exohydrolase I (1-FEH I): a vacuolar enzyme derived from a cell-wall invertase ancestor? Mass fingerprint of the 1-FEH I enzyme. Plant J 24:447–456

    Article  PubMed  Google Scholar 

  • Van den Ende W, Michiels A, Van Wonterghem D, Clerens SP, De Roover J, Van Laere AJ (2001) Defoliation induces fructan 1-exohydrolase II in witloof chicory roots cloning and purification of two isoforms, fructan 1-exohydrolase IIa and fructan 1-exohydrolase IIb mass fingerprint of the fructan 1-exohydrolase II enzymes. Plant Physiol 126:1186–1195

    Article  PubMed Central  PubMed  Google Scholar 

  • Van den Ende W, Michiels A, De Roover J, Van Laere A (2002) Fructan biosynthetic and breakdown enzymes in dicots evolved from different invertases: expression of fructan genes throughout chicory development. Sci World J 2:1273–1287

    Article  Google Scholar 

  • Van den Ende W, Clerens S, Vergauwen R, Van Riet L, Van Laere A, Yoshida M, Kawakami A (2003a) Fructan 1-exohydrolases β-(2,1)-trimmers during graminan biosynthesis in stems of wheat? Purification, characterization, mass mapping, and cloning of two fructan 1-exohydrolase isoforms. Plant Physiol 131:621–631

    Article  PubMed Central  Google Scholar 

  • Van den Ende W, De Coninck B, Clerens S, Vergauwen R, Van Laere A (2003b) Unexpected presence of fructan 6-exohydrolases (6-FEHs) in non-fructan plants: characterization, cloning, mass mapping and functional analysis of a novel “cell-wall invertase-like” specific 6-FEH from sugar beet (Beta vulgaris L.). Plant J 36:697–710

    Article  PubMed  Google Scholar 

  • Van den Ende W, Yoshida M, Clerens S, Vergauwen R, Kawakami A (2005) Cloning, characterization and functional analysis of novel 6-kestose exohydrolases (6-KEHs) from wheat (Triticum aestivum). New Phytol 166:917–932

    Article  PubMed  Google Scholar 

  • Van den Ende W, Lammens W, Van Laere A, Schroeven L, Le Roy K (2009) Donor and acceptor substrate selectivity among plant glycoside hydrolase family 32 enzymes. FEBS J 276:5788–5798

    Article  PubMed  Google Scholar 

  • Van Laere A, Van den Ende W (2002) Inulin metabolism in dicots: chicory as a model system. Plant Cell Environ 25:803–813

    Article  Google Scholar 

  • Van Riet L, Nagaraj V, Van den Ende W, Clerens S, Wiemken A, Van Laere A (2006) Purification, cloning and functional characterization of a fructan 6-exohydrolase from wheat (Triticum aestivum L.). J Exp Bot 57:213–223

    Article  PubMed  Google Scholar 

  • Van Riet L, Altenbach D, Vergauwen R, Clerens S, Kawakami A, Yoshida M, Van den Ende W, Wiemken A, Van Laere A (2008) Purification, cloning and functional differences of a third fructan 1-exohydrolase (1-FEHw3) from wheat (Triticum aestivum L.). Physiol Plant 133:242–253

    Article  PubMed  Google Scholar 

  • Verhaest M, Lammens W, Le Roy K, De Ranter CJ, Van Laere A, Rabijns A, Van den Ende W (2007) Insights into the fine architecture of the active site of chicory fructan 1-exohydrolase: 1-kestose as substrate vs sucrose as inhibitor. New Phytol 174:90–100

    Article  CAS  PubMed  Google Scholar 

  • Vitale A, Hinz G (2005) Sorting of proteins to storage vacuoles: how many mechanisms? Trends Plant Sci 10:316–323

    Article  CAS  PubMed  Google Scholar 

  • Vogt L, Ramasamy U, Meyer D, Pullens G, Venema K, Faas MM, Schols HA, de Vos P (2013) Immune modulation by different types of β2 → 1-fructans is toll-like receptor dependent. PLoS One 8:e68367

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Volenec JJ (1986) Nonstructural carbohydrates in stem base components of tall fescue during regrowth. Crop Sci 26:122–127

    Article  CAS  Google Scholar 

  • Wagner W, Wiemken A (1986) Properties and subcellular localization of fructan hydrolase in the leaves of barley (Hordeum vulgare L. cv Gerbel). J Plant Physiol 123:429–439

    Article  CAS  Google Scholar 

  • Wagner W, Wiemken A (1989) Fructan metabolism in expanded primary leaves of barley (Hordeum vulgare L. cv Gerbel): change upon ageing and spatial organization along the leaf blade. J Plant Physiol 134:237–242

    Article  CAS  Google Scholar 

  • Winter H, Robinson D, Heldt H (1993) Subcellular volumes and metabolite concentrations in barley leaves. Planta 49:180–190

    Google Scholar 

  • Xu J, Chua NH (2011) Processing bodies and plant development. Curr Opin Plant Biol 14:88–93

    Article  CAS  PubMed  Google Scholar 

  • Yamamoto S, Mino Y (1989) Mechanism of phleinase induction in stem base of orchardgrass after defoliation. J Plant Physiol 69:258–260

    Article  Google Scholar 

  • Zhang J, Dell B, Conocono E, Waters I, Setter T, Appels R (2009) Water deficit in wheat: fructan exohydrolase (1-FEH) mRNA expression and relationship to soluble carbohydrate concentration in two varieties. New Phytol 181:843–850

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by FWO (Fonds voor Wetenschappelijk Onderzoek Vlaanderen, Brussels, Belgium), the MESR (Ministère de l’Enseignement Supérieur et de la Recherche, France) [Doctoral fellowship to J.L.], UCBN (Université de Caen Basse-Normandie, France), INRA (Institut National de la Recherche Agronomique), the PHC (Programme Hubert Curien) Tournesol France-Belgium [11515WD].

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The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Institut de Recherche en Horticulture et Semences (INRA, Agrocampus-Ouest, Université d’Angers), Université d’Angers, SFR 149 QUASAV, 49045, Angers, France

    Jérémy Lothier

  2. Laboratory of Molecular Plant Biology, KU Leuven, Kasteelpark Arenberg 31, 3001, Louvain, Belgium

    André Van Laere & Wim Van den Ende

  3. Normandie Univ, Caen, France

    Marie-Pascale Prud’homme & Annette Morvan-Bertrand

  4. UMR 950 Ecophysiologie Végétale and Agronomie, Nutritions NCS Université de Caen Basse-Normandide UCBN, Esplanade de La Paix, 14032, Caen, France

    Marie-Pascale Prud’homme & Annette Morvan-Bertrand

  5. INRA, UMR 950 EVA, 14032, Caen, France

    Marie-Pascale Prud’homme & Annette Morvan-Bertrand

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  1. Jérémy Lothier
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  2. André Van Laere
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  3. Marie-Pascale Prud’homme
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  4. Wim Van den Ende
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  5. Annette Morvan-Bertrand
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Correspondence to Annette Morvan-Bertrand.

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Lothier, J., Van Laere, A., Prud’homme, MP. et al. Cloning and characterization of a novel fructan 6-exohydrolase strongly inhibited by sucrose in Lolium perenne . Planta 240, 629–643 (2014). https://doi.org/10.1007/s00425-014-2110-6

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