Biologia plantarum 46:89-93, 2003 | DOI: 10.1023/A:1022318302374
Changes in Sugar Content and Activities of Sucrose Metabolizing Enzymes in Roots and Nodules of Lentil
- 1 Department of Biochemistry, Punjab Agricultural University, Ludhiana-, India
Activities of acid and alkaline invertases and sucrose synthase were determined in roots and nodules of lentil at various stages of development. Alkaline invertase and sucrose synthase were both involved in sucrose metabolism in the nodule cytosol, but there was only a small amount of acid invertase present. Activity of sucrose metabolizing enzymes in roots was significantly less than that observed in the nodules. Amongst sugars, sucrose was found to be the main component in the host cytosol. Lentil neutral invertase (LNI) was partially purified from nodules at 50 days after sowing (DAS). Two forms of invertase were identified, i.e., a major form of 71 kDa which was taken for enzyme characterization and a minor form of 270 kDa which was not used for further studies. The purified enzyme exhibited typical hyperbolic saturation kinetics for sucrose hydrolysis. It had a Km of 11.0 to 14.0 mM for sucrose depending upon the temperature, a pH optimum of 6.8 and an optimum temperature of 40 °C. Compared with raffinose and stachyose, sucrose was better substrate for LNI. The enzyme showed no significant hydrolysis of maltose and p-nitrophenyl-α-D-glucopyranoside, showing its true β-fructosidase nature. LNI is completely inhibited by HgCl2, MnCl2 and iodoacetamide but not by CaCl2, MgCl2 or BaCl2.
Keywords: invertase; Lens culinaris; sucrose metabolism; sucrose synthase
Subjects: bacteria; invertase; Lens culinaris; lentil, sucrose metabolising enzymes; nodules, sucrose metabolising enzymes; root, sucrose metabolising enzymes; sucrose metabolising enzymes; sucrose synthase
Prepublished online: March 1, 2003; Published: July 1, 2003 Show citation
References
- Anthon, G.E., Emerich, D.W.: Developmental regulation of enzymes of sucrose and hexose metabolism in effective and ineffective soybean nodules.-Plant Physiol. 92: 346-351, 1990.
Go to original source... - Asthir, B., Singh, R.: Purification and characterization of neutral invertase from chickpea nodules.-Indian J. Biochem. Biophys. 34: 529-534, 1997.
- Awan, M.F.M.: Ontogenic variation of nodulation, nitrogen fixation and nitrogen assimilation in lentil (Lens Culinaris Medic). VI. Nitrogenase, nitrate reductase and glutamine synthetase activities.-Acta Physiol. Plant. 16: 255-262, 1994.
- Chen, J.Q., Black, C.C.: Biochemical and immunological properties of alkaline invertase isolated from sprouting soybean hypocotyls.-Arch. Biochem. Biophys. 295: 61-69, 1992.
Go to original source... - Chopra, J., Kaur, N., Gupta, A.K.: Carbohydrate status and sucrose metabolism in mungbean roots and nodules.-Phytochemistry 49: 1891-1895, 1998.
Go to original source... - Copeland, L., Lee, H.S., Cowlishaw, N.: Carbon metabolism in chickpea nodules.-Soil Biol. Biochem. 27: 381-385, 1995.
Go to original source... - Copeland, L., Vella, J., Hong, Z.: Enzymes of carbohydrate metabolism in soybean nodules.-Phytochemistry 28: 57-61, 1989.
Go to original source... - Gallagher, J.A., Pollock, C.J.: Isolation and characterization of a cDNA clone from Lolium temulentum L. encoding for a sucrose hydrolytic enzyme which shows alkaline/neutral invertase activity.-J. exp. Bot. 49: 789-795, 1998.
Go to original source... - Gascon, S., Lampen, J.O.: Purification of internal invertase of yeast.-J. biol. Chem. 243: 1567-1572, 1968.
Go to original source... - Gordon, A.J., Minchin, F.R., James, C.L., Komina, O.: Sucrose synthase in legume nodules is essential for nitrogen fixation.-Plant Physiol. 120: 867-877, 1999.
Go to original source... - Gordon, A.J., Minchin, F.R., Skot, L., James, C.L.: Stress-induced declines in soybean N2 fixation are related to nodule sucrose synthase activity.-Plant Physiol. 114: 937-946, 1997.
Go to original source... - Kouchi, H., Fukai, A., Katagiri, H., Minamisawa, K., Tajima, S.: Isolation and enzymological characterization of infected and uninfected cell protoplasts from root nodules of Glycine max.-Physiol. Plant. 73: 327-334, 1988.
Go to original source... - Lee, H., Sturm, A.: Purification and characterization of neutral and alkaline invertase from carrot.-Plant Physiol. 112: 1513-1522, 1996.
Go to original source... - Morell, M., Copeland, L.: Enzymes of sucrose breakdown in soybean nodules.-Plant Physiol. 74: 1030-1034, 1984.
Go to original source... - Morell, M., Copeland, L.: Sucrose synthase of soybean nodules.-Plant Physiol. 78: 149-154, 1985.
Go to original source... - Nelson, N.: A photometric adaptation of Somogyi method for determination of glucose.-J. biol. Chem. 153: 375-380, 1944.
Go to original source... - Ranwala, A.P., Iwanami, S.S., Masuda, H.: Acid and neutral invertases in the mesocarp of developing muskmelon Cucumis melo L. cv. Prince fruit.-Plant Physiol. 96: 881-886, 1991.
Go to original source... - Ross, H.A., Mc Rae, D., Davies, H.V.: Sucrolytic enzyme activities in cotyledons of the faba bean. Developmental changes and purification of alkaline invertase.-Plant Physiol. 111: 329-338, 1996.
Go to original source... - Sawhney, V., Sheoran, I.S., Amarjit, Singh, R.: Ontogenic changes in enzymes of sugar metabolism in relation to nitrogen fixation in chickpea (Cicer arietinum L.).-Plant Physiol. Biochem. (India) 15: 49-57, 1988.
- Singh, R., Karamdeep, L., Bhullar, S.S., Gupta, A.K.: Metabolism of free sugars in relation to the activities of enzymes involved in sucrose metabolism and nitrogen assimilation in the developing nodules of chickpea.-Plant Physiol. Biochem. 32: 875-882, 1994.
- Singh, R., Sidhu, P.S., Vadhera, S., Sital, J.S., Bhatia, I.S.: Extracellular invertase of Rhizobium japonicum and its role in free sugar metabolism in the developing root nodules of Sesbania grandiflora.-Physiol. Plant. 48: 504-508, 1980.
Go to original source... - Sukalo, V.D., Krugova, E.D.: [Enzymes of sucrose metabolism in pea nodule fractions incubated by an active strain of nodule bacteria.]-Fiziol. Biokhim. kul't. Rast. 25: 581-587, 1993. [In Russ.]
- Van den Ende, W., Van Laere, A.: Purification and properties of a neutral invertase from roots of Cichorium intybus.-Physiol. Plant. 93: 241-248, 1995.
Go to original source... - Vorster, D.J., Botha, F.C.: Partial purification and characterization of sugarcane neutral invertase.-Phytochemistry 49: 651-655, 1998.
Go to original source... - Williard, L.M., Slattery, M.: The colorimetric determination of fructosan in plant material.-J. biol. Chem. 157: 161-167, 1945.
Go to original source...



