The purification and properties of human liver ketohexokinase. A role for ketohexokinase and fructose-bisphosphate aldolase in the metabolic production of oxalate from xylitol

Abstract

Ketohexokinase (EC 2.7.1.3) was purified to homogeneity from human liver, and fructose-bisphosphate aldolase (EC 4.1.2.13) was partially purified from the same source. Ketohexokinase was shown, by column chromatography and polyacrylamide-gel electrophoresis, to be a dimer of Mr 75000. Inhibition studies with p-chloromercuribenzoate and N-ethylmaleimide indicate that ketohexokinase contains thiol groups, which are required for full activity. With D-xylulose as substrate, ketohexokinase and aldolase can catalyse a reaction sequence which forms glycolaldehyde, a known precursor of oxalate. The distribution of both enzymes in human tissues indicates that this reaction sequence occurs mainly in the liver, to a lesser extent in the kidney, and very little in heart, brain and muscle. The kinetic properties of ketohexokinase show that this enzyme can phosphorylate D-xylulose as readily as D-fructose, except that higher concentrations of D-xylulose are required. The kinetic properties of aldolase show that the enzyme has a higher affinity for D-xylulose 1-phosphate than for D-fructose 1-phosphate. These findings support a role for ketohexokinase and aldolase in the formation of glycolaldehyde. The effect of various metabolites on the activity of the two enzymes was tested to determine the conditions that favour the formation of glycolaldehyde from xylitol. The results indicate that few of these metabolites affect the activity of ketohexokinase, but that aldolase can be inhibited by several phosphorylated compounds. This work suggests that, although the formation of oxalate from xylitol is normally a minor pathway, under certain conditions of increased xylitol metabolism oxalate production can become significant and may result in oxalosis.

Properties of Normal and Mutant Recombinant Human Ketohexokinases and Implications for the Pathogenesis of Essential Fructosuria

“…This is low, compared with 18 (16) or 12 units/mg (20) for the rat enzyme and 17 units/mg for the bovine enzyme (1). The higher activity of the recombinant human enzyme may therefore indicate some loss of activity in the human liver enzyme obtained by Bais et al (2). Consistent with this, we do observe variability in specific activity of recombinant KHK-C, the highest value for any one freshly purified preparation being 15.5 units/mg (k cat ϭ 8.4 s Ϫ1 ).…”

“…The properties of recombinant KHK-C purified in the present study are consistent with those of purified hepatic enzymes (1,2,16,20). Our second finding, that the KHK-A isoform is indeed biochemically active, suggests that the low levels of KHK-A expression seen in a wide range of tissues probably do fulfill some specific physiologic function.…”

“…A number of other furanose sugars can also act as KHK substrates (1). KHK activity is highest in the liver, followed by renal cortex and small intestine (2). Its primary role in these sites seems to be clearance of dietary fructose through a specialized pathway involving aldolase B and triokinase.…”

Inhibition of Nicotinamide Phosphoribosyltransferase (NAMPT), an Enzyme Essential for NAD+ Biosynthesis, Leads to Altered Carbohydrate Metabolism in Cancer Cells

“…3, 4A, and 5). Fourth, the absence of detectable levels of fructose and sedoheptulose in the FK866-treated cells (data not shown) ruled out a direct phosphorylation event involving enzymes like ketohexokinase and sedoheptulokinase (33,34). In addition, the expression of ketohexokinase is diminished in the human clear cell type of renal cell carcinoma (35).…”

Gene and protein expression changes in response to normoxic perfusion in mouse hearts

“…Khk expression had a significant decline after 60 and also 120 min of perfusion. This enzyme is expressed predominantly in the liver, to a lesser extent in the kidney, and very little in heart, brain and muscle (Bais, James, Rofe, & Conyers, 1985).…”