| Allosteric regulation is an essential mechanism utilized by all organisms to control a vast array of biochemical reactions. Despite interest in this field for over 100 years, the mechanism of long-range allosteric regulation is still poorly understood, particularly for cases in which structural changes are minimal. Recent studies have identified an important role for protein motions in several allosteric enzymes. It is unknown whether this is a generalizable trend. To gain improved understanding of how allosteric signals are transduced across protein-protein interfaces, the enzymes imidazole glycerol phosphate synthase (IGPS) and ribonuclease Z (tRNase Z) have been studied. In each system, an emphasis has been placed on characterizing the ternary complex in relation to the effector and substrate bound states to allow for the identification of elements essential for allostery.;Studies of IGPS have identified a central role for protein motions in allosteric regulation. Millisecond motions for backbone and side chain resonances increase substantially upon binding of the effector molecule, PRFAR, as evidenced by relaxation dispersion and severe resonance broadening. Comparisons between the effector bound state with the ternary complex has identified a reduced subset of residues that form an unbroken network connecting the effector binding site with the protein-protein interface. Global analysis of these motions provides evidence of an additional state not represented by one of the four that have been studied (apo, substrate bound (acivicin-glutamine analog), effector bound (PRFAR) and ternary). These results are contrary to the Monod-Wyman-Changeux model of protein allostery, which has gained popularity in recent years.;Preliminary studies of tRNase Z have allowed for the assignment of 53% of backbone resonances. Initial characterization of protein motions has identified increased flexibility for an arm domain involved in substrate recognition and proper cleavage and elevated transverse relaxation rates due to conformational exchange (Rex) for residues implicated in the coordination of catalytic Zn2+ ions. An assay and several point mutants of conserved residues have been designed which will allow for future mechanistic studies of the allosteric regulation in tRNase Z. |
