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Gluconeogenic Fructose 1, 6-bisphosphatase from Ginkgo biloba L.: Isolation and characterization

Sachina Yonzone, Suchisree Jha, Dhani Raj Chhetri*

Abstract


Fructose 1, 6-bisphostase (FBPase) was partially purified from mature leaves of Ginkgo biloba which is a rare medicinal plant effective as antioxidant, adaptogenic and memory enhancer. The purification procedure involved homogenization, low speed centrifugation, streptomycin sulphate precipitation, ammonium sulphate fractionation and chromatography on Bio Gel-A 0.5 m. The level of purification was about 22 fold and the recovery was about 56% at the final stage of purification. D-fructose-1,6-bisphosphate (FBP) was found to be the specific substrate for Ginkgo biloba FBPase. Among the other hexose phosphates, D-glucose-6-phosphate, D-fructose-6-phosphate and D-galactose-6-phosphate could not act as substrates for this enzyme. However, this enzyme showed traces of activity in presence of D-fructose-1-phosphate. With the increase in substrate (FBP) concentration from 0 to 8 mM the activity of the enzyme increased linearly. The FBPase activity was directly proportional to the time of incubation upto 60 minutes and with respect to protein concentration upto 400 µg. The Bio Gel A 0.5 m purified G. biloba FBPase showed a Km values of 1.86 mM for D-fructose-1,6-bisphosphate. The FBPase required Mg2+ as a cofactor. Mn2+ and Mg2+ appreciably stimulated the FBPase activity in a concentration guided manner. The monovalent cations, K+ and Na+ had little effect while Zn2+ was extremely inhibitory to the enzyme.

Keywords


Ginkgo biloba; enzyme purification; enzyme characterization; fructose 1,6-bisphosphatase; D-fructose-1,6-bisphosphate; gluconeogenesis

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References


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