Structural elements in IGP synthase exclude water to optimize ammonia transfer.
In the complex pathway of histidine biosynthesis, a key branch point linking amino acid and purine biosynthesis is catalyzed by the bifunctional enzyme imidazole glycerol phosphate (IGP) synthase. The first domain of IGP synthase, a triad glutamine amidotransferase, hydrolyzes glutamine to form glutamate and ammonia. Its activity is tightly regulated by the binding of the substrate PRFAR to its partner synthase domain. Recent crystal structures and molecular dynamics simulations strongly suggest that the synthase domain, a (beta/alpha)(8) barrel protein, mediates the insertion of ammonia and ring formation in IGP by channeling ammonia from one remote active site to the other. Here, we combine both mutagenesis experiments and computational investigations to gain insight into the transfer of ammonia and the mechanism of conduction. We discover an alternate route for the entrance of ammonia into the (beta/alpha)(8) barrel and argue that water acts as both agonist and antagonist to the enzymatic function. Our results indicate that the architecture of the two subdomains, most notably the strict conservation of key residues at the interface and within the (beta/alpha)(8) barrel, has been optimized to allow the efficient passage of ammonia, and not water, between the two remote active sites.
Amaro
Rommie E
RE
Department of Chemistry, University of Illinois, Urbana, Illinois, USA.
Myers
Rebecca S
RS
Davisson
V Jo
VJ
Luthey-Schulten
Zaida A
ZA
D000595:Amino Acid Sequence; D000619:Aminohydrolases / Q000737:chemistry*; D000641:Ammonia / Q000737:chemistry*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D005972:Glutaminase / Q000737:chemistry; D006639:Histidine / Q000737:chemistry; D007700:Kinetics; D008958:Models, Molecular; D015233:Models, Statistical; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010649:Phenylalanine / Q000737:chemistry; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity; D020124:Thermotoga maritima / Q000201:enzymology; D014867:Water / Q000737:chemistry
D014867:Water; D010649:Phenylalanine; D006639:Histidine; D000641:Ammonia; C087227:imidazole glycerol phosphate synthase; D005972:Glutaminase; D000619:Aminohydrolases