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Partial purification and characterisation of cytosolic Fructose-1, 6-bisphosphatase from Drymaria cordata

Niki Doma Sherpa, Raksha Mukhia, Dhani Raj Chhetri

Abstract


Drymaria cordata is an important ethnomedicinal plant from which many important secondary metabolites have been reported. Partial purification of the enzyme, fructose 1,6-bisphosphatase was carried out following the methods of homogenization, streptomycin sulphate precipitation, ammonium sulphate cut and molecular sieve chromatography through Bio-Gel A-0.5m column. Biochemical characterization experiments were performed by standard methods with the enzyme preparation as purified from the column. Cytosolic fructose 1,6-bisphosphatase from the leaves of Drymaria cordata was purified to about 27-fold with 77% of recovery over homogenate fraction. The enzyme was highly specific to D-fructose-1,6-bisphosphate. With increase of protein concentration upto 300µg and incubation time upto120 minutes, the enzyme activity increased linearly. The metal ions Mg2+ or Mn2+ strongly stimulated the enzyme activity on the other hand Li+, Hg2+ and Zn2+ were potent inhibitors. The D. cordata enzyme showed temperature maxima at 40˚C while the optimum pH was at 8.0. The Km value of the enzyme for its substrate, Fructose 1,6-bisphosphate was 1.11µM proving its strong affinity.

Keywords


Fructose 1,6-bisphosphatase, Fructose 1,6-bisphosphate, Drymaria cordata, protein purification, enzyme characterization

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References


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DOI: https://doi.org/10.21746/aps.2018.7.4.9

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