Synthesis of L-2-(N-Pteroylamino )-3-(N -phosphonoacetyl)aminopropanoic Acid as an Analogue of the Putative Phosphorylated Intermediate in the γ-Glutamation of Folic Acid by Folylpolyglutamate Synthetase

Abstract

L-2-(N-Pteroyl)amino-3-(N-phosphonoacetyl)aminopropanoic acid was synthesized as an analogue of the putative y-phosphorylated intermediate in the enzyme-catalyzed γ-glutamation of folic acid by folylpolyglutamate synthetase (FPGS). N-(Benzyloxycarbonyl)-L-aspartic acid was converted in four steps to methyl L-2-(N-benzyloxycarbonyl)amino-3-aminopropanoate, and the latter was allowed to react with p-nitrophenyl dimethoxyphosphonoacetate to obtain methyl L-2-(N-benzyloxycarbonylamino)- 3-(N-dimethoxyphosphonoacetyl)aminopropanoate. After catalytic hydrogenation, the resulting amine was coupled to N10-formylpteroic acid via the mixed carboxylic-carbonic anhydride method, and the three ester groups were removed by sequential treatment with Me3SiBr in DMF and NaOH in DMSO. When the last step was performed only with NaOH/DMSO, one of the phosphonate esters remained intact, giving L-2-(N -pteroyl )amino-3 -(N -monOInethoxyphosphonoacetyl )aminopropanoic acid. Also synthesized as a potential FPGS inhibitor was Nα-(4-amino-4-deoxy-N10-methylpteroyl)-Nε-phosphonoacetyl- L-Iysine. The ability of these phosphonoacetyl derivatives to inhibit catalytic addition of L-glutamic acid to folic acid proved to be very low, suggesting that replacement of the CH2C(=O)OP(=O)(OH)2 moiety by NHC(=O)CH2P(=O)(OH)2 may place the terminal phosphonyl group in an unfavorable spatial orientation for binding to the enzyme.

Keywords

• folic acid
• folylpolyglutamate synthetase
• γ- phosphorylated intermediate analogue