Aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/IMP cyclohydrolase (ATIC) is a bifunctional enzyme with folate-dependent AICAR transformylase and IMP cyclohydrolase activities that catalyzes the last two steps of purine biosynthesis. The AICAR transformylase inhibitors BW1540 and BW2315 are sulfamido-bridged 5,8-dideazafolate analogs with remarkably potent K(i) values of 8 and 6 nm, respectively, compared with most other antifolates. Crystal structures of ATIC at 2.55 and 2.60 A with each inhibitor, in the presence of substrate AICAR, revealed that the sulfonyl groups dominate inhibitor binding and orientation through interaction with the proposed oxyanion hole. These agents then appear to mimic the anionic transition state and now implicate Asn(431') in the reaction mechanism along with previously identified key catalytic residues Lys(266) and His(267). Potent and selective inhibition of the AICAR transformylase active site, compared with other folate-dependent enzymes, should therefore be pursued by further design of sulfonyl-containing antifolates.
Within de novo purine biosynthesis, the AICAR transformylase and IMP cyclohydrolase activities of the bifunctional enzyme ATIC convert the intermediate AICAR to the final product of the pathway, IMP. Identification of the AICAR transformylase active site and a proposed formyl transfer mechanism have already resulted from analysis of crystal structures of avian ATIC in complex with substrate and/or inhibitors. Herein, we focus on the IMPCH active site and the cyclohydrolase mechanism through comparison of crystal structures of XMP inhibitor complexes of human ATIC at 1.9 A resolution with the previously determined avian enzyme. This first human ATIC structure was also determined to ascertain whether any subtle structural differences, compared to the homologous avian enzyme, should be taken into account for structure-based inhibitor design. These structural comparisons, as well as comparative analyses with other IMP and XMP binding proteins, have enabled a catalytic mechanism to be formulated. The primary role of the IMPCH active site appears to be to induce a reconfiguration of the substrate FAICAR to a less energetically favorable, but more reactive, conformer. Backbone (Arg64 and Lys66) and side chain interactions (Thr67) in the IMPCH active site reorient the 4-carboxamide from the preferred conformer that binds to the AICAR Tfase active site to one that promotes intramolecular cyclization. Other backbone amides (Ile126 and Gly127) create an oxyanion hole that helps orient the formyl group for nucleophilic attack by the 4-carboxamide amine and then stabilize the anionic intermediate. Several other residues, including Lys66, Tyr104, Asp125, and Lys137', provide substrate specificity and likely enhance the catalytic rate through contributions to acid-base catalysis.
Evidence
2:
Inferred from Physical InteractionUniProtKB
Aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/IMP cyclohydrolase (ATIC) is a bifunctional enzyme with folate-dependent AICAR transformylase and IMP cyclohydrolase activities that catalyzes the last two steps of purine biosynthesis. The AICAR transformylase inhibitors BW1540 and BW2315 are sulfamido-bridged 5,8-dideazafolate analogs with remarkably potent K(i) values of 8 and 6 nm, respectively, compared with most other antifolates. Crystal structures of ATIC at 2.55 and 2.60 A with each inhibitor, in the presence of substrate AICAR, revealed that the sulfonyl groups dominate inhibitor binding and orientation through interaction with the proposed oxyanion hole. These agents then appear to mimic the anionic transition state and now implicate Asn(431') in the reaction mechanism along with previously identified key catalytic residues Lys(266) and His(267). Potent and selective inhibition of the AICAR transformylase active site, compared with other folate-dependent enzymes, should therefore be pursued by further design of sulfonyl-containing antifolates.
The chemical reactions and pathways resulting in the formation of IMP, inosine monophosphate, by the stepwise assembly of a purine ring on ribose 5-phosphate.
The progression of the brainstem from its formation to the mature structure. The brainstem is the part of the brain that connects the brain with the spinal cord.
The process whose specific outcome is the progression of the cerebellum over time, from its formation to the mature structure. The cerebellum is the portion of the brain in the back of the head between the cerebrum and the pons. In mice, the cerebellum controls balance for walking and standing, modulates the force and range of movement and is involved in the learning of motor skills.
The progression of the cerebral cortex over time from its initial formation until its mature state. The cerebral cortex is the outer layered region of the telencephalon.
J. Biol. Chem. 271, 2225-2233 (1996)[PubMed:8567683]
We report here the cloning and sequencing of the cDNA, purification, steady state kinetic analysis, and truncation mapping studies of the human 5-aminoimidazole- 4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (AICARFT/IMPCHase). These steps of de novo purine biosynthesis, respectively. In all species of both prokaryotes and eukaryotes studied, these two activities are present on a single bifunctional polypeptide encoded on the purH gene. The human purH cDNA is 1776 base pairs in length encoding for a 591-amino acid polypeptic (Mr = 64,425). The human and avian purH cDNAs are 75 and 81% similar on the nucleotide and amino acid sequence level, respectively. The Km values for AICAR and (6R,6S)10-formyltetrahydrofolate are 16.8 microM +/- 1.5 and 60.2 microM +/- 5.0, respectively, for the cloned, purified human enzyme. A 10-amino acid sequence within the COOH-terminal portion of human AICARFT/IMPCHase has some degree of homology to a previously noted "folate binding site." Site directed mutagenesis studies indicate that this sequence plays no role in enzymatic activity. We have constructed truncation mutants which demonstrate that each of the two enzyme activities can be expressed independent of the other. IMPCHase and AICARFT activities are located within the NH2-terminal 223 and COOH-terminal 406 amino acids, respectively. The truncation mutant possessing AICARFT activity displays steady state kinetic parameters identical to those of the holoenzyme.
The chemical reactions and pathways involving a nucleoside, a nucleobase linked to either beta-D-ribofuranose (a ribonucleoside) or 2-deoxy-beta-D-ribofuranose, (a deoxyribonucleoside), e.g. adenosine, guanosine, inosine, cytidine, uridine and deoxyadenosine, deoxyguanosine, deoxycytidine and thymidine (= deoxythymidine).
Any process that results in a change in state or activity of a cell or an organism (in terms of movement, secretion, enzyme production, gene expression, etc.) as a result of an inorganic substance stimulus.
The chemical reactions and pathways resulting in the formation of tetrahydrofolate, 5,6,7,8-tetrahydrofolic acid, a folate derivative bearing additional hydrogens on the pterin group.
IEAOrtholog Compara
Enzymatic activity
This protein acts as an enzyme. It is known to catalyze the following reactions
Protein involved in the biosynthesis of purine, a nitrogenous heterocyclic base, e.g. adenine, guanine, hypoxanthine and xanthine. De novo synthesis involves a complex, energy-expensive pathway that yields inosine 5'-monophosphate (IMP), a purine ribonucleotide. AMP and GMP are then formed from IMP in separate pathways. Adenine and guanine are found in both DNA and RNA. Hypoxanthine and xanthine are important intermediates in the synthesis and degradation of the purine nucleotides.
Enzyme which catalyzes hydrolysis reaction, i.e. the addition of the hydrogen and hydroxyl ions of water to a molecule with its consequent splitting into two or more simpler molecules.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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