Fructose 3-phosphate, a novel monosaccharide phosphate, has been identified in the lens of diabetic rats. This compound, which is not present in normal lenses, is a protein glycosylating agent and enzyme inactivator. In addition, because of its structural features, this metabolite is relatively labile and undergoes hydrolysis to yield inorganic phosphate and the potent glycosylating agent, 3-deoxyglucosone. The increase in the concentration of fructose 3-phosphate in the lens of diabetic rats suggests that it and its hydrolysis product, 3-deoxyglucosone, may be responsible in part for the development of some diabetic complications in the lens.

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In human erythrocytes, the first step in the metabolism of fructose is generally thought to be phosphorylation to fructose 6-phosphate catalysed by hexokinase. In variance with this assumption, we show here that fructose in these cells is metabolized primarily to fructose 3-phosphate by a specific 3-phosphokinase. This process has an overall estimated Km of 30 mM with respect to extracellular fructose and an apparent Vmax. of 0.6 mumol/h per ml. At a fixed concentration of fructose in the medium, the accumulation of fructose 3-phosphate was linearly dependent on the duration of incubation up to 5 h and was not affected by glucose. Once accumulated, fructose 3-phosphate appears to be degraded and/or relatively slowly metabolized, decreasing by only approximately 30% after a 12 h incubation in a fructose-free medium.

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The title compounds (14a,b) were 5' epimers of a derivative of a phosphonate isostere of ATP in which the CH2OP alpha system of ATP was replaced by CH(R)CH2P alpha [R = L-S(CH2)2CH(NH2)CO2H]. They resisted synthesis via attempted S-alkylation of the corresponding epimeric 5'-mercapto derivatives. A practicable route to 14a,b commenced with Michael condensation of L-homocysteine with the diphenyl ester of the 5',6'-vinyl phosphonate analogue of 2',3'-O-isopropylideneadenosine 5'-phosphate. The resulting epimeric 5' thioethers were separated by reverse-phase HPLC. The two phenyl groups were replaced by benzyl groups, after which the alpha-amino acid residue was protected as an N-Boc methyl ester. Both benzyl groups were removed by hydrogenolysis, and the resulting phosphonic acid was converted into its pyrophosphoryl derivative. Blocking groups were then removed under conditions that furnished 14a and 14b without racemization of their L-amino acid residues. Also synthesized were the P beta-NH-P gamma imido analogue (15a) of 14a and the sulfoxide derivative (16a) of 14a. The structures of 14a and 16a were verified by FAB mass spectra, which revealed the protonated molecular ions of their sodium salts. All adducts appeared to function as dual substrate site inhibitors (competitive to ATP and to methionine) of the rat normal tissue (MAT-2) form of methionine adenosyltransferase (MAT); 14a and 15a [KM(ATP)/Ki = 4 and 9, respectively] were the most effective. Adduct 15a was the most effective inhibitor [KM(ATP)/Ki = 13] of the MAT-T form from rat hepatoma tissue; the kinetic data indicated dual-site inhibition by 15a with apparently complete coverage of the ATP site and incomplete coverage of the methionine site. The inhibition properties of the adducts indicated little preference in the order in which the two MAT forms bound ATP and methionine.