Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Transferase other than ribonuclease
Reexamination Certificate
1997-05-09
2001-04-03
Achutamurthy, Ponnathapu (Department: 1652)
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Transferase other than ribonuclease
C536S023200
Reexamination Certificate
active
06210943
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Technical Field
The present invention relates to a sucrose phosphate synthase from Citrus including its isoform, and to DNA encoding the enzyme.
2. Earlier Technology
Sucrose is a transport form of the photoassimilate in most plants. The sucrose in mature leaves (source) is mostly transported by the phloem to the plant organs (sink) that are net consumers of the photo-assimilate. A key enzyme of sucrose synthesis pathway, sucrose-phosphate synthase (EC 2.4.1.14) (hereinafter referred to as “SPS”), catalyzes the following reaction:
Fructose 6-phosphate+UDP glucose→Sucrose 6-phosphate+UDP Sucrose 6-phosphate is converted to sucrose by sucrose phosphatase.
Considerable interest has focused on the role of SPS in regulation of sucrose synthesis in source leaves (Kerr, P. S. and Huber, S. C. (1987) Planta 170:197-204; and Echeverria, E. and Burns, J. K. (1989) Plant Physiol. 90:530-533). SPS activity itself has been found in many plants, for example cucurbits (Lingle, S. E. and Dunlap, J. R. (1987) Plant Physiol. 84:386-389; Hubbard, N. L. et al. (1989) Plant Physiol. 91:1527-1534; and Burger, Y. and Schaffer, A. A. (1991) Sucrose metabolism in mature fruit peduncles of
Cucumis melo
and
Cucumis sativus.
In:Recent advances in phloemtransport and assimilate partitioning, pp. 244-247, Bonnemain, J. L. et al. eds. ouest Editions, Nantes, France), peach (Hubbard, N. L. et al. (1991) Physiol. Plant 82:191-196), pear (Moriguchi, T. et al. (1992) J. Am. Soc. Hortic. Sci. 117:247-278), and celery (Stoop, J. M. H. and Pharr, D. M. (1994) J. Am. Soc. Hortic. Sci. 119:237-242), sugar beet (Fieuw, S. and Willenbrink, J. (1987) J. Plant Physiol. 131:153-162), sugar cane (Wendler, R. et al. (1990) Planta 183:31-39; and Goldner, W. et al. (1991) Plant Sci. 73:143-147), sucrose-accumulating Lycopersicon spp. (Miron, D. and Schaffer, A. A. (1991) Humb. And Bonpl. Plant Physiol. 95:623-627), Dali, N. et al. (1992) Plant Physiol. 99:434-438; and Stommel, J. R. (1992) Plant Physiol. 99:324-328), rice (Smyth, D. A. and Prescott, H. E. (1989) Plant Physiol. 89:893-896), strawberry (Hubbard, N. L. et al. (1991) Physiol. Plant 82:191-196), and citrus (Lowell, C. A. et al. (1989) Plant Physiol. 90:1394-1402; and Echeverria, E. (1992) Plant Sci. 85:125-129).
To investigate the enzymatic function of SPS on sucrose biosynthesis, SPS has been purified to near homogeneity from spinach (Salvucci, M.E. et al. (1990) Arch. Biochem. Biophys. 281:212-218), wheat (Salerno, G. L. et al. (1991) Physiol. Plant 81:541-547), and maize (Bruneau, J. -M. et al. (1991) Plant Physiol. 96:473-478). SPS is an allosteric enzyme which is activated by binding of the substrate-similar glucose-6-phosphate and inhibited by P
i
at the allosteric site (Doehlert, D. C. and Huber, S. C. (1983) Plant Physiol. 73:989-994). In addition, the activity of SPS is regulated by protein phosphorylation (Huber, J. L. A. et al. (1989) Arch. Biochem. Biophys. 270:681-690; Siegl, G. et al. (1990) FEBS Letters 270:198-202; and Huber, S. C. and Huber J. L. (1991) Plant Cell Physiol. 32:319-326). Recently, the function and structure of SPS have also been studied at the molecular level in maize (Worrell, A. C. et al. (1991) Plant Cell 3:1121-1130), spinach (Klein, R. R. et al. (1993) Planta 190:498-510), and sugar beet (Hesse, H. et al. (1995) Mol. Gen. Genet. 247:515-520).
In citrus, the sucrose accumulation is one of the very important events in fruit development. Phloem-free juice sacs at the middle stage of fruit development showed higher SPS activity than the adjacent transport tissues, vascular nodules and segment epidermis (Lowell, C. A. et al. (1989) Plant Physiol. 90:1394-1402). However, analysis of the function and expression of SPS at the molecular level has been quite limited in Citrus.
An object of this invention is to clone cDNA for SPS from Citrus and characterize it at the molecular level.
Another object of the invention is to provide an SPS from Citrus.
SUMMARY OF THE INVENTION
This invention provides DNA encoding a sucrose phosphate synthase from Citrus having an amino acid sequence shown in SEQ ID NO:2, or an isoform thereof sharing at least 50% homology with said sucrose phosphate synthase in amino acid level.
In a preferred embodiment of the invention, the isoform is a different type of sucrose phosphate synthase from Citrus, containing a partial amino acid sequence shown in SEQ ID NO:4 or SEQ ID NO:5.
In another embodiment, the DNA has a nucleotide sequence shown in SEQ ID NO:1 that encodes the mature form of the sucrose phosphate synthase.
This invention further provides a sucrose phosphate synthase from Citrus or an isoform thereof, as defined above.
REFERENCES:
Lee, C.C. et al., Science, vol. 239, pp. 1288-1291, 1988.*
Ann C. Worrell et al., “Expresseion of a Maize Sucrose Phosphate Synthase in Tomato Alters Leaf Carbohydrate Partitioning,” The Plant Cell, vol. 3, pp. 1121-1130, Oct. 1991.
Robert R. Klein et al, “Cloning and Developmental Expression of the Sucrose-Phosphate-Synthase Gene from Spinach,” Planta, vol. 190, No. 4, pp. 498-510, 1993.
Holger Hesse et al., “Cloning and Expression Analysis of Sucrose-Phosphate Synthase from Sugar Beet (Beta Vulgaris L.),” Mol Gen Genet, vol. 247, pp. 515-520, 1995.
A. Komatsu et al., Programme and Abstracts, VIII Congress of the International Society of Citriculture, May 1996, P039, p. 97.
Akira Komatsu et al., “Cloning and Molecular Analysis of cDNAs Encoding Three Sucrose Phosphate Synthase Isoforms from a Citrus Fruit (Citrus Unshiu Marc.),” Mol Gen Genet, vol. 252, pp. 346-351, 1996.
Akira Komatsu, “Molecular and Physiological Characterization for Sucrose-Metabolizing Enzymes and Genes in Sink Tissues of Citrus,” pp. 1-97, 1996. Thesis, Meiji University.
Achutamurthy Ponnathapu
Nissan Chemical Industries Ltd.
Oblon & Spivak, McClelland, Maier & Neustadt P.C.
Tung Peter P.
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