Allosteric pathways in imidazole glycerol phosphate synthase

Abstract

Protein allosteric pathways are investigated in the imidazole glycerol phosphate synthase heterodimer in an effort to elucidate how the effector (PRFAR,formimino]-5-aminoimidazole-4-carboxamide ribonucleotide) activates glutaminase catalysis at a distance of 25 Å from the glutamine-binding site. We apply solution NMR techniques and community analysis of dynamical networks, based on mutual information of correlated protein motions in the active and inactive enzymes. We find evidence that the allosteric pathways in the PRFAR bound enzyme involve conserved residues that correlate motion of the PRFAR binding loop to motion at the protein-protein interface, and ultimately at the glutaminase active site. The imidazole glycerol phosphate synthase bienzyme is an important branch point for the histidine and nucleotide biosynthetic pathways and represents a potential therapeutic target against microbes. The proposed allosteric mechanism and the underlying allosteric pathways provide fundamental insights for the design of new allosteric drugs and/or alternative herbicides.glutamine hydrolysis | protein networks | generalized correlation analysis | network theory A llostery is a fundamental property that allows for the regulation of function and dynamic adaptability of enzymes and proteins. Allosteric enzymes contain at least two distant binding sites, including the active site responsible for catalytic activity, which binds the substrate, and the allosteric site, which binds the effector and initiates the allosteric signal propagation to the active site. In V-type systems, substrate binding is not affected by the presence of the effector but if the effector is not bound, the allosteric protein is usually catalytically inactive (or poorly active), indicating that the effector binding is coupled to the kinetic and/or thermodynamic parameters of the biochemical reaction in the active site. Allosteric information transfer can range from large, enthalpically driven conformational changes to purely entropically driven motions or a combination of both enthalpic and entropic effects, but in each case the kinetic parameters of the catalyzed reaction at the substrate binding site are altered. At the heart of allosterism there is intramolecular thermodynamic coupling over long distances (>10 Å), between the active and allosteric sites. An important challenge for fundamental studies is the elucidation of the allosteric pathways that connect the two ligand-binding sites.In this work, we combine community network analysis based on molecular dynamics (MD) simulations and NMR studies of protein motion based on relaxation dispersion techniques and chemical shift titrations experiments to provide an atomistic description of allostery in the V-type allosteric enzyme imidazole glycerol phosphate synthase (IGPS) from the thermophile Thermotoga maritima (Fig. 1). IGPS is a tightly associated heterodimeric enzyme in which each monomer enzyme catalyzes a different reaction (1-3). The 23 kDa HisH enzyme is a member of the glutamine amidotransferas...

Allosteric networks in thrombin distinguish procoagulant vs. anticoagulant activities

“…Allosteric networks were characterized using a community network analysis approach previously applied to investigate allostery in tRNA-protein complexes and other protein systems (24,31,32). This approach constructs a dynamic contact map consisting of a network graph in which each residue is treated as a "node" connected by edges to other nodes when two residues are deemed to be "in contact."…”

Dynamical network of residue–residue contacts reveals coupled allosteric effects in recognition, catalysis, and mutation

“…Another analysis that involves monitoring the time evolution of residue-residue contact formation and breaking has been proven useful in identifying certain characteristic events during conformational transitions (25). Correlated motions between different biomolecular segments have been identified using cross-correlation analysis and building dynamical networks (25)(26)(27)(28)(29). However, interpretation of the results of such analysis in case of CypA, which exhibit subtle changes upon substrate binding and catalysis, has been ambiguous.…”

Altering the allosteric pathway in IGPS suppresses millisecond motions and catalytic activity

“…Recently, we have advanced these studies and demonstrated that activation of IGPS is dependent on the ability of allosteric effectors to stimulate domainwide motions in the HisF subunit and ligands that induce greater motion within the central (αβ) 8 barrel are more effective activators of glutaminase chemistry (22). NMR and computational community network studies proposed an optimal allosteric pathway through networks of HisF amino acid residues that link the PRFAR binding site and adjacent flexible loop 1 to residues comprising a central hydrophobic cluster essential for catalysis, as well as salt bridge residues at the dimer interface (25,26). These previous results suggest important amino acids in each community that, upon mutation, should disrupt the IGPS allosteric network.…”

“…(25) In particular, there was an increase in communication between the HisF community, f3′, which contained secondary-structure elements fβ1, fβ2, fβ3, fβ4, fα2, fα2, and hα1, and community h2′, which encompassed the glutaminase active site and hP10 (SI Appendix, Fig. S1).…”