The kinetic mechanism of 5-enolpyruvylshikimate-3-phosphate synthase from a Gram-positive pathogen Streptococcus pneumonia

The Streptococcus pneumoniae 5-enolpyruvylshikimate-3-phosphate (EPSP) synt hase is a potential novel antibacterial target. The enzyme catalyzes a reve rsible transfer of an enolpyruvyl group from phospho(enol)pyruvate (PEP) to shikimate 3-phosphate (S3P) to give EPSP with the release of inorganic pho sphate (Pi). Understanding the kinetic mechanism of this enzyme is crucial to the design of novel inhibitors of this enzyme that may have potential as antibacterial agents. Steady-state kinetic studies of product inhibition a nd inhibition by glyphosate (GLP) have demonstrated diverse inhibition patt erns of the enzyme. In the forward reaction, GLP is a competitive inhibitor with respect to PEP, but an uncompetitive inhibitor relative to S3P. Produ ct inhibition shows that EPSP is a competitive inhibitor versus both PEP an d S3P, suggesting that the forward reaction follows a random sequential mec hanism. In the reverse reaction, GLP is an uncompetitive inhibitor versus E PSP, but a noncompetitive inhibitor versus Pi. This indicates that a non-pr oductive quaternary complex might be formed between the enzyme, EPSP, GLP a nd Pi. Product inhibition in the reverse reaction has also been investigate d. The inhibition patterns of the S. pneumoniae EPSP synthase are nor entir ely consistent with those of EPSP synthases from other species, indicating that EPSP synthases from different organisms may adopt unique mechanisms to catalyze the same reactions.