Allosteric Inhibition of ATP Phosphoribosyltransferase by Protein:Dipeptide and Protein:Protein Interactions.

ATP phosphoribosyltransferase (ATPPRT) catalyzes the first step of histidine biosynthesis in bacteria, namely, the condensation of ATP and 5-phospho-α-d-ribosyl-1-pyrophosphate (PRPP) to generate -(5-phospho-β-d-ribosyl)-ATP (PRATP) and pyrophosphate. Catalytic (HisG) and regulatory (HisZ) subunits assemble in a hetero-octamer where HisZ activates HisG and mediates allosteric inhibition by histidine. In , HisG is necessary for the bacterium to persist in the lung during pneumonia. Inhibition of ATPPRT is thus a promising strategy for specific antibiotic development. Here, ATPPRT is shown to follow a rapid equilibrium random kinetic mechanism, unlike any other ATPPRT. Histidine noncompetitively inhibits ATPPRT. Binding kinetics indicates histidine binds to free ATPPRT and to ATPPRT:PRPP and ATPPRT:ATP binary complexes with similar affinity following a two-step binding mechanism, but with distinct kinetic partition of the initial enzyme:inhibitor complex. The dipeptide histidine-proline inhibits ATPPRT competitively and likely uncompetitively, respectively, against PRPP and ATP. Rapid kinetics analysis shows His-Pro binds to the ATPPRT:ATP complex via a two-step binding mechanism. A related HisZ that shares 43% sequence identity with HisZ is a tight-binding allosteric inhibitor of HisG. These findings lay the foundation for inhibitor design against ATPPRT.