INVERSION OF AN ASYMMETRIC CENTER IN CARBOCYCLIC INHIBITORS OF 3-DEHYDROQUINATE SYNTHASE - EXAMINING AND EXPLOITING THE MECHANISM FOR SYN-ELIMINATION DURING SUBSTRATE TURNOVER

Conversion of 3-deoxy-D-arabino-heptulosonic acid 7-phosphate (DAHP) i nto 3-dehydroquinate (DHQ) by the enzyme DHQ synthase has been propose d to proceed through a step where the phosphate monoester of a reactiv e intermediate mediates its own elimination. This hypothesis was teste d by challenging DHQ synthase with a series of carbocyclic substrate a nalogues possessing an inverted methine carbon relative to the same as ymmetric center in substrate DAHP which loses a proton during eliminat ion of inorganic phosphate. Despite the stereochemical alteration, epi carbocyclic substrate analogues 5-[(phosphonooxy)methyl]-5-deoxyquinat e, 5-(phosphonomethyl)-5-deoxyquinate, 5-(phosphonoethyl)-5-deoxyquina te, and 3-(phosphonooxy)quinate inhibited DHQ synthase with respective inhibition constants (K-i) of 30 nM, 55 nM, 30 mu M, and 53 mu M. The se inhibitors were synthesized from quinic acid and with the exception of 3-(phosphono oxy)quinate, were assembled via a strategy employing intramolecular, radical cyclization to establish the stereocenter wher e the (phosphonooxy)methyl, phosphonomethyl, and phosphonoethyl moieti es were attached to the carbocyclic ring. The observed diastereomeric promiscuity in the binding of epicarbocyclic substrate analogues by DH Q synthase is consistent with the hypothesized nonenzymic syn-eliminat ion of inorganic phosphate during substrate turnover.