New EPSP synthase inhibitors: Synthesis and evaluation of an aromatic tetrahedral intermediate mimic containing a 3-malonate ether as a 3-phosphate surrogate

A new analog of the EPSP synthase enzyme reaction intermediate 1, containing a 3-malonate ether moiety in place of the usual 3-phosphate group, was synthesized from 3,5-dihydroxybenzoic acid. This simple, synthetically accessible aromatic compound (5) is an effective competitive inhibitor versus S3P with an apparent K 1 of 1.3 ± 0.22 μM. This result demonstrates that a simple benzene ring can be a suitable achiral substitute for the more complex shikimate ring in the design of EPSP synthase inhibitors. Furthermore, the greater potency of 5 versus the phenol 6, glycolate 7 and the gallic acid analog 8 demonstrates the requirement for multiple anionic charges at the dihydroxybenzoate 5-position in order to attain effective inhibition of this enzyme. However, this 3-malonate ether substituted compound was at least 10-fold less effective as a bisubstrate inhibitor than the corresponding 3-phosphate. This suggests that tetrahedral intermediate mimics possessing a 3-malonate ether moiety are less effective than their corresponding 3-phosphates in accessing the optimal enzyme conformation stabilizing 1. Aromatic analogues of the EPSP synthase reaction substrate, product, and tetrahedral intermediate were synthesized from 3,5-dihydroxybenzoic acid, containing a 3-malonate ether in place of the normal 3-phosphate group. These molecules more clearly define the scope and limitations of incorporating 3-malonate ethers as 3-phosphate replacements in this system. The potency of 5 suggests that a benzene ring is an effective substitute for the more complex shikimate ring in EPSP synthase inhibitors.