Study On The Gene-Expression Analysis And Enzymatic Properties Of Isocitrate Dehydrogenase From Ectomycorrhiza Of Pinus Sylvestris Var. Mongolica Litv

Isocitrate dehydrogenase(IDH) is an important rate-limiting enzyme in the TCA cycle. It is the best model to study both the function of proteins and the catalytic mechanism and regulatory mechanism of enzyme. The IDH of mycorrhizal tissue, root tissue and fungus mycelium were purified by physical methods. The relative expression of IDH was studied by real-time fluorescent quantitative. The experiment provides a theoretical basis for the study on the relationship between function and metabolism of IDH in ectomycorrhiza.IDH were purified by physical methods and tested by SDS-PAGE electrophoresis. SDS-PAGE showed that purified IDH gave a single band. The IDHs subunit relative molecular mass of IDH from mycorrhizal tissue, root tissue and fungus mycelium were about 46kD. The kinetic study showed that the Km for NADP+ of mycorrhizal tissue, root tissue and fungus mycelium were 10.7μM/L,11.4μM/L and 22.1μM/L. The Km for isocitrate of mycorrhizal tissue, root tissue and fungus mycelium were 71.7μM/L,79.3μM/L and 87.8μM/L. The optimal pH of mycorrhizal tissue, root tissue and fungus mycelium were 8.2,8.0 and 7.5. The optimal temperatures were 45℃,45℃ and 42℃, respectively. The activities of the three IDHs were dependent on the binding of divalent metal ions. The maximum activities of IDHs were observed with the presence of Mn2+. The results showed that relative expression of the IDH gene in mycorrhizal tissue was higher than that in root tissue. Based on the test results we can conclude that ectomycorrhizal fungi can greatly improve the activity and gene expression of IDH of host plants. The results provide the basis for biochemistry and molecular biology to further explanation how the ectomycorrhiza improves the growth condition of host plants.