Purification and properties of liver fructose 1,6-bisphosphatase from C57BL/KsJ normal and diabetic mice.

Fructose 1,6-bisphosphatase was purified from the livers of C57BL/KsJ mice to apparent homogeneity.The enzyme was shown to be a tetramer of a subunit M, = -35,000.The purified enzyme showed maximal activity at neutral pH; high affinity for its substrate, fructose 1,6-bisphosphate; requirement for a divalent cation (M inhibition by AMP; susceptibility to limited proteolysis by subtilisin; and activation by K+ or m+.Thallium and lithium, two potential NMR probes to study the enzyme-monovalent metal interactions, affected fructose 1,6-bisphosphatase activity.Thallium was found to be an activator.V,, with T I + equals that obtained with K+, but the affinity of the enzyme for T l ' (KO = 1 6 ~) was about 3 times greater than for K'.Lithium was strongly inhibitory.The apparent Ki values for Li' were 0.8 and 0.3 m~ in the presence of saturating concentrations of M g + and Mn2+, respectively.Mouse liver fructose 1,g-bisphosphatase, either in its native state or in 1.6 M urea, was not a substrate for the catalytic subunit of cyclic AMPdependent protein kinase.This is in contrast with the results obtained with rat liver fructose 1,6-bisphosphatase.The rat liver enzyme was phosphorylated by the above kinase and the same maximal phosphate incorporation (about 1 mol of phosphate/mol of enzyme protomer) was obtained in either the presence or absence of urea.Fructose 1,6-bisphosphatase was found to be increased in the livers of the genetically diabetic mouse (CS'IBL/KsJ-db strain).The enzyme from these animals was also purified to homogeneity.No difference in either molecular or enzymatic properties was found between the two purified enzymes.