Plant Molecular Biology

, Volume 58, Issue 2, pp 213–227 | Cite as

Essential amino acids of starch synthase IIa differentiate amylopectin structure and starch quality between japonica and indica rice varieties

  • Yasunori Nakamura
  • Perigio B. Francisco
  • Yuko Hosaka
  • Aya Sato
  • Takayuki Sawada
  • Akiko Kubo
  • Naoko Fujita


Four amino acids were variable between the ‘active’ indica-type and ‘inactive’ japonica-type soluble starch synthase Ha (SSIIa) of rice plants; Glu-88 and Gly-604 in SSIIa of indica-cultivars IR36 and Kasalath were replaced by Asp-88 and Ser-604, respectively, in both japonica cultivars Nipponbare and Kinmaze SSIIa, whereas Val-737 and Leu-781 in indica SSIIa were replaced by Met-737 in cv. Nipponbare and Phe-781 in cv. Kinmaze SSIIa, respectively. The SSIIa gene fragments shuffling experiments revealed that Val-737 and Leu-781 are essential not only for the optimal SSIIa activity, but also for the capacity to synthesize indica-type amylopectin. Surprisingly, however, a combination of Phe-781 and Gly-604 could restore about 44% of the SSIIa activity provided that Val-737 was conserved. The introduction of the ‘active’ indica-type SSIIa gene enabled the japonica-type cv. Kinmaze to synthesize indica-type amylopectin. The starch in the transformed japonica rice plants exhibited gelatinization-resistant properties that are characteristic of indica-rice starch. Transformed lines expressing different levels of the IR36 SSIIa protein produced a variety of starches with amylopectin chain-length distribution patterns that correlated well with their onset temperatures of gelatinization. The present study confirmed that the SSIIa activity determines the type of amylopectin structure of rice starch to be either the typical indica-type or japonica-type, by playing a specific role in the synthesis of the long B1 chains by elongating short A and B1 chains, notwithstanding the presence of functional two additional SSII genes, a single SSI gene, two SSIII genes, and two SSIV genes in rice plants.

Key words

amylopectin indica rice japonica rice starch starch synthase transgenic rice 



degree of polymerization


differential scanning calorimetry


granule-bound starch synthase




souluble starch synthase


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  1. Ball, S.G. and Morell, M.K. 2003. From bacterial glycogen to starch: understanding the biogenesis of the plant starch granule Ann. Rev. Plant Biol. 54: 207–233.CrossRefGoogle Scholar
  2. Buschiazzo, A., Ugalde, J.E., Guerin, M.E., Shepard, W., Ugalde, R.A. and Alzari, P.M. 2004. Crystal structure of glycogen synthase: homologous enzymes catalyze glycogen synthesis and degradation EMBO J. 23: 3196–3205.PubMedCrossRefGoogle Scholar
  3. Cao, H., Imparl-Radosevich, J., Guan, H., Keeling, P.L., James, M.G. and Myers, A.M. 1999. Identification of the soluble starch synthase activities of maize endosperm Plant Physiol. 120: 205–215.PubMedCrossRefGoogle Scholar
  4. Commuri, P.D. and Keeling, P.L. 2001. Chain-length specificities of maize starch synthase I enzyme: studies of glucan affinity and catalytic properties Plant J. 25: 475–486.PubMedCrossRefGoogle Scholar
  5. Craig, J., Lloyd, J.R., Tomlinson, K., Barber, L., Edwards, A., Wang, T.L., Martin, C., Hedley, C.L. and Smith, A.M. 1998. Mutations in the gene encoding starch synthase II profoundly alter amylopectin structure in the pea embryos Plant Cell 10: 413–426.PubMedCrossRefGoogle Scholar
  6. Dian, W., Jiang, H. and Wu, P. 2005. Evolution and expression analysis of starch synthase III and IV J. Exp. Bot. 56: 623–632.PubMedCrossRefGoogle Scholar
  7. Edwards, A., Fulton, D.C., Hylton, CM., Jobling, S.A., Gidley, M., Rössner, U., Martin, C. and Smith, A.M. 1999. A combined reduction in activity of starch synthase II and III of potato has novel effects on the starch of tubers Plant J. 17: 251–261.CrossRefGoogle Scholar
  8. Furukawa, K., Tagaya, M., Tanizawa, K. and Fukui, T. 1994. Identification of Lys277 at the active site of Escherichia coli glycogen synthase. Application of affinity labeling combined with site-directed mutagenesis J. Biol. Chem. 269: 868–871.PubMedGoogle Scholar
  9. Gallant, D.J., Bouchet, B. and Baldwin, P.M. 1997. Microscopy of starch: evidence of a new level of granule organization Carbohydr. Polym. 32: 177–191.CrossRefGoogle Scholar
  10. Gao, Z., Keeling, P.L., Shibles, R. and Guan, H. 2004. Involvement of lysine-193 of the conserved “K-T-G-G” motif in the catalysis of maize starch synthase Ha Arch. Biochem. Biophys. 427: 1–7.PubMedCrossRefGoogle Scholar
  11. Gao, M., Wanat, J., Stinard, P.S., James, M.G. and Myers, A.M. 1998. Characterization of dull1, a maize gene coding for a novel starch synthase Plant Cell 10: 399–412.PubMedCrossRefGoogle Scholar
  12. Harn, C., Knight, M., Ramakrishnan, A., Guan, H., Keeling, P.L. and Wasserman, B.P. 1998. Isolation and characterization of the zSSIIa and zSSIIb starch synthase cDNA clones from maize endosperm Plant Mol. Biol. 37: 639–649.PubMedCrossRefGoogle Scholar
  13. Hirose, T. and Terao, T. 2004. A comprehensive expression analysis of the starch synthase gene family in rice (Oryza sativa L.) Planta 220: 9–16.PubMedCrossRefGoogle Scholar
  14. Hood, E.E., Gelvin, S.B., Melchers, L.S. and Hoekema, A. 1993. New Agrobacterium helper plasmids for gene transfer to plants Transgenic Res. 2: 208–218.CrossRefGoogle Scholar
  15. Horibata, T., Nakamoto, M., Fuwa, H. and Inouchi, N. 2004. Structural and physicochemical characteristics of endosperm starches of rice cultivars recently bred in Japan J. Appl. Glycosci. 51: 303–313.Google Scholar
  16. Imparl-Radosevich, J.M., Keeling, P.L. and Guan, H. 1999. Essential arginine residues in maize starch synthase IIa are involved in both ADP-glucose and primer binding FEBS Lett. 457: 357–362.PubMedCrossRefGoogle Scholar
  17. James, M.G., Denyer, K. and Myers, A.M. 2003. Starch synthesis in the cereal endosperm Curr. Opin. Plant Biol. 6: 215–222.PubMedCrossRefGoogle Scholar
  18. Jiang, H., Dian, W., Liu, F. and Wu, P. 2004. Molecular cloning and expression analysis of three genes encoding starch synthase II in rice Planta 218: 1062–1070.PubMedCrossRefGoogle Scholar
  19. Jobling, S. 2004. Improving starch for food and industrial applications Curr. Opin. Plant Biol. 7: 210–218.PubMedCrossRefGoogle Scholar
  20. Kubo, A., Rahman, S., Utsumi, Y., Li, Z., Mukai, Y., Yamamoto, M., Ugaki, M., Harada, K., Satoh, H., Konik-Rose, C., Morell, M. and Nakamura, Y. 2005. Complementation of sugary-1 phenotype in rice endosperm with the wheat Isoamylasel gene supports a direct role for isoamylasel in amylopectin biosynthesis Plant Physiol. 137: 43–56.PubMedCrossRefGoogle Scholar
  21. Morell, M.K., Kosar-Hashemi, B., Cmiel, M., Samuel, M.S., Chandler, P., Rahman, S., Buleon, A., Batey, I.L. and Li, Z. 2003. Barley sex6 mutants lack starch synthase IIa activity and contain a starch with novel properties Plant J. 34: 173–185.PubMedCrossRefGoogle Scholar
  22. Mu-Forster, C, Huang, R., Powers, J.R., Harriman, R.W., Knight, M., Singletary, G.W., Keeling, P.L. and Wasserman, B. 1996. Physical association of starch biosyntheitic enzymes with starch granules of maize endosperm Plant Physiol. 111: 821–829.PubMedCrossRefGoogle Scholar
  23. Nakamura, Y. 2002. Towards a better understanding of the metabolic system for amylopectin biosynthesis in plants: rice endosperm as a model tissue Plant Cell Physiol. 43: 718–725.PubMedCrossRefGoogle Scholar
  24. Nakamura, Y., Sakurai, A., Inaba, Y., Kimura, K., Iwasawa, N. and Nagamine, T. 2002. The fine structure of amylopectin in endosperm from Asian cultivated rice can be largely classified into two classes Starch 54: 117–131.CrossRefGoogle Scholar
  25. Nichols, D.J., Keeling, P., Spalding, M. and Guan, H. 2000. Involvement of conserved aspartate and glutamate residues in the catalysis and substrate binding of maize starch synthase Biochemistry 39: 7820–7825.PubMedCrossRefGoogle Scholar
  26. Nishi, A., Nakamura, Y., Tanaka, N. and Satoh, H. 2001. Biochemical and genetic analysis of the effects of amylose-extender mutation in rice endosperm Plant Physiol. 127: 459–472.PubMedCrossRefGoogle Scholar
  27. Oka, H. and Morishima, H. 1997. Wild and cultivated rice. In: T. Matsuo, Y. Futsuhara, F. Kikuchi and H. Yamaguchi (Eds.), Science of the Rice Plants. Genetics, Nobunkyo, Tokyo, 88–111 pp.Google Scholar
  28. Shannon, J.C. and Garwood, D.L. 1984. In: Starch: Chemistry and Technology, Academic Press, Orlando, FL, 25–86 pp.Google Scholar
  29. Smith, A.M., Denyer, K. and Martin, C. 1997. The synthesis of the starch granules Ann. Rev. Plant Physiol. Plant Mol. Biol. 48: 67–87.CrossRefGoogle Scholar
  30. Takaiwa, F., Yamanouchi, U., Yoshihara, T., Washida, H., Tanabe, F., Kato, A. and Yamada, K. 1996. Characterization of common cis-regulatory elements responsible for the endosperm-specific expression of members of the rice glutelin Plant Mol. Biol. 30: 1207–1221.PubMedCrossRefGoogle Scholar
  31. Takeda, Y. and Hizukuri, S. 1987. Structures of rice amylopectins with low and high affinities for iodine Carbohydr. Res. 168: 79–88.CrossRefGoogle Scholar
  32. Tetlow, I.J., Wait, R., Lu, Z., Akkasaeng, R., Bowsher, C.G., Esposito, S., Kosar-Hashemi, B., Morell, M.K. and Emes, M.J. 2004. Protein phosphorylation in amyloplasts regulates starch branching enzyme activity and protein-protein interactions Plant Cell 16: 694–708.PubMedCrossRefGoogle Scholar
  33. Thompson, D.B. 2000. On the non-random nature of amylopectin branching Carbohydr. Polym. 43: 223–239.CrossRefGoogle Scholar
  34. Umemoto, T., Aoki, N., Lin, H., Nakamura, Y., Inouchi, N., Sato, Y., Yano, M., Hirabayashi, H. and Maruyama, S. 2004. Natural variation in rice starch synthase IIa affects enzyme and starch properties Functional Plant Biol. 31: 671–684.CrossRefGoogle Scholar
  35. Umemoto, T., Nakamura, Y., Satoh, H. and Terashima, K. 1999. Differences in amylopectin structure between two rice varieties in relation to the effects of temperature during growth Starch 51: 58–62.CrossRefGoogle Scholar
  36. Umemoto, T., Yano, M., Satoh, H., Shomura, A. and Nakamura, Y. 2002. Mapping of a gene responsible for the difference in amylopectin structure between japonica-type and indica-type rice varieties Theor. Appl. Genet. 104: 1–8.PubMedCrossRefGoogle Scholar
  37. Warth, F.J. and Darabsett, D.B. 1914. Disintegration of rice grains by means of alkari Bull. Agric. Res. Inst. Pasa 38: 1–9.Google Scholar
  38. Wong, K.S., Kubo, A., Jane, J.L., Harada, K., Satoh, H. and Nakamura, Y. 2003. Structures and properties of amylopectin and phytoglycogen in the endosperm of sugary-1 mutant of rice J. Cereal Sci. 37: 139–149.CrossRefGoogle Scholar
  39. Wu, C.Y., Suzuki, A., Washida, H. and Takaiwa, F. 1998. The GCN4 motif in a rice glutelin gene is essential for endosperm-specific gene expression and is activated by Opaque-2 in transgenic rice plants Plant J. 14: 673–683.PubMedCrossRefGoogle Scholar
  40. Yamamori, M., Fujita, S., Hayakawa, K., Matsuki, J. and Yasui, T. 2000. Genetic elimination of a starch granule protein, SGP-1, of wheat generates an altered starch with apparent high amylase Theor. Appl. Genet. 101: 21–29.CrossRefGoogle Scholar
  41. Yep, A., Ballicora, M.A., Sivak, M.N. and Preiss, J. 2004. Identification and characterization of a critical region in the glycogen synthase from Escherichia coli J. Biol. Chem. 279: 8359–8367.PubMedCrossRefGoogle Scholar
  42. Zhang, X., Colleoni, C, Ratushna, V., Sirghie-Colleoni, M., James, M.G. and Myers, A.M. 2004. Molecular characterization demonstrates that the Zea mays gene sugary2 codes for the starch synthase isoform SSIIa Plant Mol. Biol. 54: 865–879.PubMedCrossRefGoogle Scholar
  43. Zhenyu, G., Dali, Z., Xia, C., Yihua, Z., Meixian, Y., Danian, H., Jiayang, L. and Qian, Q. 2003. Map-based cloning of the ALK gene, which controls the gelatinization temperature of rice Sci. China 46: 661–668.CrossRefGoogle Scholar

Copyright information

© Springer 2005

Authors and Affiliations

  • Yasunori Nakamura
    • 1
    • 2
  • Perigio B. Francisco
    • 2
  • Yuko Hosaka
    • 2
  • Aya Sato
    • 2
  • Takayuki Sawada
    • 2
  • Akiko Kubo
    • 2
  • Naoko Fujita
    • 1
    • 2
  1. 1.Akita Prefectural UniversityAkita-CityJapan
  2. 2.CRESTJapan Science and Technology AgencyKawaguchi, SaitamaJapan

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