Partial purification and characterization of Hexokinase from developing grains of thermotolerant wheat (Triticum aestivum L. em. Thell)”

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dc.contributor.advisorSingal, H. R.
dc.contributor.authorFageria, Leena
dc.date.accessioned2016-09-14T10:39:22Z
dc.date.available2016-09-14T10:39:22Z
dc.date.issued2013
dc.description.abstractWheat grain is the dominant grain of world commerce and is the staple food of millions of people worldwide. High temperature beyond 30 0 C, which is usually encountered during later part of grain filling period, affects grain yield (reduction by 20-50 per cent) and grain quality. Starch is the major storage carbohydrate in wheat grains. It is synthesized from sucrose, which is the principal product of leaf photosynthesis and transported to the wheat grain. As this sucrose enters the cell, it is metabolized to produce fructose and glucose. Hexokinase (HXK) is an important enzyme as it catalyzes the irreversible phosphorylation of hexoses to hexose-phosphates. The hexose-phosphates produced catalyzes synthesis of starch. Keeping above in view, the present investigations were conducted to purify and characterize hexokinase from developing grains of thermotolerant wheat. Hexokinase was purified to near homogeneity (as revealed by single band on Native-PAGE) from immature grains (21 days after anthesis) of thermotolerant wheat WH-1021 by using conventional protein purification techniques viz. 30-60% ammonium sulphate fractionation, DEAE-cellulose ion exchange chromatography and gel filtration through sephadex G-100. The enzyme was purified about 19.27 fold with 48.43 per cent recovery. The molecular weight as determined by gel filtration and subunit molecular weight as determined by SDS-PAGE (single band) were found to be 50 kDa, indicating that enzyme is a monomer. The purified enzyme exhibited optimum activity at pH 8 and 35 0 C. It was thermostable upto 50 0 C. The enzyme followed Michaelis-Menten kinetics with Km value of 2.5 mM and 1.82 mM for Glc and ATP as substrate, respectively. Amongst various nucleotides and metabolites tested AMP, ADP, UDP, CTP and G-6-P were found to be the potent inhibitors of purified hexokinase, inhibiting the enzyme activity by 13, 16, 5, 37 and 13%, respectively. However, GTP, UTP, G-1,6-BP and F-1,6-BP acted as poor inhibitors of the enzyme. The enzyme activity was stimulated by K + and Mg 2+ while Cu 2+ and Co 2+ inhibited the activity at 5 mM concentration. To summarize, higher thermostability of enzyme is suggestive of enzyme’s adaptation to high temperature stress.en_US
dc.identifier.urihttp://krishikosh.egranth.ac.in/handle/1/76586
dc.language.isoenen_US
dc.publisherCCSHAUen_US
dc.subBiochemistry
dc.subjectfruits, vegetables, insecticides, extraction, biological interaction, yields, organic amendments, garlic, livestock, biological phenomenaen_US
dc.these.typeM.Sc
dc.titlePartial purification and characterization of Hexokinase from developing grains of thermotolerant wheat (Triticum aestivum L. em. Thell)”en_US
dc.typeThesisen_US
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Partial purification and characterization of Hexokinase from developing grains of thermotolerant wheat (Triticum aestivum L. em. Thell)”
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