Cereal Research Communications
Volume/Issue:
Volume 37: Issue 3
Isolation and expression of a new high molecular weight glutenin subunit gene at the Glu-D-1-2 locus from Aegilops tauschii
Authors: Y. Zhang, X. An 1 , X. Li 1 , S. Chen 1 , L. Gao 1 , K. Wang 1 , S. Wang 1 and Y. Yan 1
View More View Less
  • 1 Capital Normal University Key Laboratory of Genetics and Biotechnology, College of Life Science Beijing 100048 China
  • 2 College of Applied Sciences and Humanities of Beijing Union University Beijing 100083 China
DOI:
https://doi.org/10.1556/crc.37.2009.3.14
Page Count:
449–457
Publication Date:
01 Sep 2009
Online Publication Date:
08 Sep 2009
Article Category:
Research Article
Restricted access

Two new y-type HMW-GSs in Ae. tauschii , 1Dy12.1* t and 1Dy12.2 t with the mobility order of 1Dy12.2 t > 1Dy12.1* t > 1Dy12.1 t >1Dy12, were identified by both SDS-PAGE and MALDI-TOF-MS. Molecular cloning and sequencing showed that the genes encoding subunits 1Dy12.1* t and 1Dy12.2 t had identical nucleotide acid sequences with 1,947 bp encoding a mature protein of 627 residues. Their deduced molecular weights were 67,347.6 Da, satisfactorily corresponding to that of 1Dy12.2 t subunit determined by MALDI-TOF-MS (67,015.7 Da), but was significantly smaller than that of the the 1Dy12.1* t subunit (68,577.1 Da). Both subunits showed high similarities to 1Dy10, suggesting that they could have a positive effect on bread-making quality. Interestingly, the expressed protein of the cloned ORF from accessions TD87 and TD130 in E. coli co-migrated with subunit 1Dy12.2 t , but moved slightly faster than 1Dy12.1* t on SDS-PAGE. The expressed protein in transgenic tobacco seeds, however, had the same mobility as the 1Dy12.1* t subunit, as confirmed by both SDS-PAGE and Western blotting. Although direct evidence of phosphoprotein could not be obtained by specific staining method, certain types of post-translational modifications (PTMs) of the 1Dy12.1* t subunit could not be excluded. We believe PTMs might be responsible for the molecular weight difference between the subunits 1Dy12.1* t and 1Dy12.2 t .

  • Agrawal, G.K., Thelen, J.J. 2005. Development of a simplified economical polyacrylamide gel staining protocol for phosphoproteins. Proteomics 5 :4684–4688.

    Thelen J.J. , 'Development of a simplified economical polyacrylamide gel staining protocol for phosphoproteins ' (2005 ) 5 Proteomics : 4684 -4688.

    • Search Google Scholar

  • Bevan, M. 1984. Binary Agrobacterium vectors for plant transformation. Nucleic Acid Res. 12 :8711–8721.

    Bevan M. , 'Binary Agrobacterium vectors for plant transformation ' (1984 ) 12 Nucleic Acid Res : 8711 -8721.

    • Search Google Scholar

  • Dong, H., Cox, T.S., Sears, R.G., Lookhart, G.L. 1991. High molecular weight glutenin genes: Effects on quality in wheat. Crop Sci. 31 :974–979.

    Lookhart G.L. , 'High molecular weight glutenin genes: Effects on quality in wheat ' (1991 ) 31 Crop Sci : 974 -979.

    • Search Google Scholar

  • Flavell, R.B., Doldsbrough, A.P., Robert, L.S., Schnick, D., Thompson, R.D. 1989. Genetic variation in wheat HMW glutenin subunits and the molecular basis of bread-making quality. Bio/Tech 7 :1281–1285.

    Thompson R.D. , 'Genetic variation in wheat HMW glutenin subunits and the molecular basis of bread-making quality ' (1989 ) 7 Bio/Tech : 1281 -1285.

    • Search Google Scholar

  • Hickman, D.R., Roepstorff, P., Shewry, P.R., Tatham, A.S. 1995. Molecular weights of high molecular weight subunits of glutenin determined by mass spectrometry. J. Cereal Sci. 22 :99–103.

    Tatham A.S. , 'Molecular weights of high molecular weight subunits of glutenin determined by mass spectrometry ' (1995 ) 22 J. Cereal Sci. : 99 -103.

    • Search Google Scholar

  • Hsam, S.L.K., Kieffer, R., Zeller, F.J. 2001. Significance of Aegilops tauschii glutenin genes on breadmaking properties of wheat. Cereal Chem. 78 :521–525.

    Zeller F.J. , 'Significance of Aegilops tauschii glutenin genes on breadmaking properties of wheat ' (2001 ) 78 Cereal Chem : 521 -525.

    • Search Google Scholar

  • Lamacchia, C., Shewry, P.R., Fonzo, N.D., Forsyth, J.L., Harris, N., Lazzeri, P.A., Napier, J.A., Halford, N.G., Barcelo, P. 2001. Endosperm-specific activity of a storage protein gene promoter in transgenic wheat seed. J. Exp. Bot. 52 :243–250.

    Barcelo P. , 'Endosperm-specific activity of a storage protein gene promoter in transgenic wheat seed ' (2001 ) 52 J. Exp. Bot. : 243 -250.

    • Search Google Scholar

  • Li, X.H., Zhang, Y.Z., Gao, L.Y., Wang, A.L., Ji, K.M., He, Z.H., Appels, R., Ma, W.J., Yan, Y.M. 2007. Molecular cloning, heterologous expression and phylogenetic analysis of a novel y-type HMW glutenin subunit gene from G genome of Triticum timopheevi . Genome 50 :1130–1140.

    Yan Y.M. , 'Molecular cloning, heterologous expression and phylogenetic analysis of a novel y-type HMW glutenin subunit gene from G genome of Triticum timopheevi ' (2007 ) 50 Genome : 1130 -1140.

    • Search Google Scholar

  • Li, X.H., Ma, W., Gao, L.Y., Zhang, Y.Z., Wang, A.L., Ji, K.M., Wang, K., Appels, R., Yan, Y. 2008. A novel chimeric LMW-GS gene from the wild relatives of wheat Ae. kotschyi and Ae. juvenalis : evolution at the Glu-3 loci. Genetics 180 :93–101.

    Yan Y. , 'A novel chimeric LMW-GS gene from the wild relatives of wheat Ae. kotschyi and Ae. juvenalis: evolution at the Glu-3 loci ' (2008 ) 180 Genetics : 93 -101.

    • Search Google Scholar

  • Matheis, G. 1999. Phosphorylation of food proteins with phosphorus oxychloride improvement of functional and nutritional properties: A review. Food Chem. 39 :13–26.

    Matheis G. , 'Phosphorylation of food proteins with phosphorus oxychloride improvement of functional and nutritional properties: A review ' (1999 ) 39 Food Chem : 13 -26.

    • Search Google Scholar

  • Payne, P.I., Corfield, K.G. 1979. Subunit composition of wheat glutenin proteins, isolated by gel filtration in a dissociating medium. Planta 145 :83–88.

    Corfield K.G. , 'Subunit composition of wheat glutenin proteins, isolated by gel filtration in a dissociating medium ' (1979 ) 145 Planta : 83 -88.

    • Search Google Scholar

  • Pfluger, L.A., D’Ovidio, R., Margiotta, B., Pena, R., Mujeeb-Kazi, A., Lafiandra, D. 2001. Characterisation of high- and low-molecular weight glutenin subunits associated to the D genome of Aegilops tauschii in a collection of synthetic hexaploid wheats. Theor. Appl. Genet. 103 :1293–1301.

    Lafiandra D. , 'Characterisation of high- and low-molecular weight glutenin subunits associated to the D genome of Aegilops tauschii in a collection of synthetic hexaploid wheats ' (2001 ) 103 Theor. Appl. Genet. : 1293 -1301.

    • Search Google Scholar

  • Robert, L.S., Thompson, R.D., Flavell, R.B. 1989. Tissue-specific expression of a wheat high molecular weight glutenin in transgenic tobacco. Plant Cell 1 :569–578.

    Flavell R.B. , 'Tissue-specific expression of a wheat high molecular weight glutenin in transgenic tobacco ' (1989 ) 1 Plant Cell : 569 -578.

    • Search Google Scholar

  • Salzano, A.M., Crescenzi, M. 2005. Mass spectrometry for protein identification and the study of post translational modifications. Ann. Ist. Super. Sanità. 41 :443–450

    Crescenzi M. , 'Mass spectrometry for protein identification and the study of post translational modifications ' (2005 ) 41 Ann. Ist. Super. Sanità. : 443 -450.

    • Search Google Scholar

  • Shewry, P.R., Halford, N.G., Tatham, A.S. 1992. The high molecular weight subunits of wheat glutenin. J. Cereal Sci. 15 :105–120.

    Tatham A.S. , 'The high molecular weight subunits of wheat glutenin ' (1992 ) 15 J. Cereal Sci. : 105 -120.

    • Search Google Scholar

  • Yan, Y., Hsam, S.L.K., Yu, J.Z., Jiang, Y., Zeller, F.J. 2003a. Allelic variation of the HMW glutenin subunits in Aegilops tauschii accessions detected by Sodium Dodecyl Sulphate (SDS-PAGE), Acid Polyacrylamide Gel (A-PAGE) and Capillary Electrophoresis. Euphytica 130 :377–385.

    Zeller F.J. , 'Allelic variation of the HMW glutenin subunits in Aegilops tauschii accessions detected by Sodium Dodecyl Sulphate (SDS-PAGE), Acid Polyacrylamide Gel (A-PAGE) and Capillary Electrophoresis ' (2003 ) 130 Euphytica : 377 -385.

    • Search Google Scholar

  • Yan, Y., Hsam, S.L.K., Yu, J.Z., Jiang, Y., Ohtsuka, I., Zeller, F.J. 2003b. HMW and LMW glutenin alleles among putative tetraploid and hexaploid European spelt wheat ( Triticum spelta L.) progenitors. Theor. Appl. Genet. 107 :1321–1330.

    Zeller F.J. , 'HMW and LMW glutenin alleles among putative tetraploid and hexaploid European spelt wheat (Triticum spelta L.) progenitors ' (2003 ) 107 Theor. Appl. Genet. : 1321 -1330.

    • Search Google Scholar

  • Yan, Y., Yu, J., Jiang, Y., Hu, Y., Cai, M., Hsam, S.L.K., Zeller, F.J., 2003c. Capillary electrophoresis separation of high molecular weight glutenin subunits in bread wheat ( Triticum aestivum L.) and related species with phosphate-based buffers. Electrophoresis 24 :1429–1436.

    Zeller F.J. , 'Capillary electrophoresis separation of high molecular weight glutenin subunits in bread wheat (Triticum aestivum L.) and related species with phosphate-based buffers ' (2003 ) 24 Electrophoresis : 1429 -1436.

    • Search Google Scholar

  • Yan, Y., Zheng, J., Xiao, Y., Yu, J., Hu, Y., Cai, M., Li, Y., Hsam, S.L.K., Zeller, F.J. 2004a. Identification and molecular characterization of a novel y-type Glu-Dt1 glutenin gene of Aegilops tauschii . Theor. Appl. Genet. 108 :1349–1358.

    Zeller F.J. , 'Identification and molecular characterization of a novel y-type Glu-Dt1 glutenin gene of Aegilops tauschii ' (2004 ) 108 Theor. Appl. Genet. : 1349 -1358.

    • Search Google Scholar

  • Yan, Y., Jiang, Y., Sun, M., Yu, J., Xiao, Y., Zheng, J., Hu, Y., Cai, M., Li, Y., Hsam, S.L.K., Zeller, F.J. 2004b. Rapid identification of HMW glutenin subunits from different hexaploid wheat species by acidic capillary electrophoresis. Cereal Chem. 81 :561–566.

    Zeller F.J. , 'Rapid identification of HMW glutenin subunits from different hexaploid wheat species by acidic capillary electrophoresis ' (2004 ) 81 Cereal Chem. : 561 -566.

    • Search Google Scholar

  • Zhang, Q., Dong, Y.M., An, X.L., Wang, A.L., Zhang, Y.Z., Li, X.H., Xia, X.C., He, Z.H., Yan, Y.M. 2008a. Characterization of HMW glutenin subunits in common heat and related species by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS). J. Cereal Sci. 47 :252–261.

    Yan Y.M. , 'Characterization of HMW glutenin subunits in common heat and related species by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) ' (2008 ) 47 J. Cereal Sci. : 252 -261.

    • Search Google Scholar

  • Zhang, Y.Z., Li, X.H., Wang, A.L., An, X.L., Zhang, Q., Pei, Y.H., Gao, L.Y., Ma, W.J., Appels, R., Yan, Y.M. 2008b. Novel x-type HMW glutenin genes from Aegilops tauschii and their implications on the wheat origin and evolution mechanism of Glu-D1-1 proteins. Genetics 178 :23–33.

    Yan Y.M. , 'Novel x-type HMW glutenin genes from Aegilops tauschii and their implications on the wheat origin and evolution mechanism of Glu-D1-1 proteins ' (2008 ) 178 Genetics : 23 -33.

    • Search Google Scholar

  • Zhang, Y.Z., Li, Q.Y., Yan, Y.M., Zheng, J.G., An, X.L., Xiao, Y.H., Wang, A.L., Pei, Y.H., Wang, H.B., Hsam, S.L.K., Zeller, F.J. 2006. Molecular characterization and phylogenetic analysis a novel glutenin gene ( Dy10.1 t ) from Aegilops tauschii . Genome 49 :735–745.

    Zeller F.J. , 'Molecular characterization and phylogenetic analysis a novel glutenin gene (Dy10.1t) from Aegilops tauschii ' (2006 ) 49 Genome : 735 -745.

    • Search Google Scholar
Cover Cereal Research Communications
Cereal Research Communications A Quarterly of the Cereal Research Non-Profit Ltd. Company
Print ISSN:
0133-3720
Online ISSN:
1788-9170
Keywords:
Aegilops tauschii ; common wheat; HMW-GS; SDS-PAGE; MALDI-TOF-MS; protein post-translational modifications