Browse
Submit Data
Databases
API
Help

Dataset Information

27 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Structural basis for the catalytic mechanism of human phosphodiesterase 9.

ABSTRACT: The phosphodiesterases (PDEs) are metal ion-dependent enzymes that regulate cellular signaling by metabolic inactivation of the ubiquitous second messengers cAMP and cGMP. In this role, the PDEs are involved in many biological and metabolic processes and are proven targets of successful drugs for the treatments of a wide range of diseases. However, because of the rapidity of the hydrolysis reaction, an experimental knowledge of the enzymatic mechanisms of the PDEs at the atomic level is still lacking. Here, we report the structures of reaction intermediates accumulated at the reaction steady state in PDE9/crystal and preserved by freeze-trapping. These structures reveal the catalytic process of a PDE and explain the substrate specificity of PDE9 in an actual reaction and the cation requirements of PDEs in general.

SUBMITTER: Liu S

PROVIDER: S-EPMC2533186 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Structural basis for the catalytic mechanism of human phosphodiesterase 9.

Liu Shenping S Mansour Mahmoud N MN Dillman Keith S KS Perez Jose R JR Danley Dennis E DE Aeed Paul A PA Simons Samuel P SP Lemotte Peter K PK Menniti Frank S FS

Proceedings of the National Academy of Sciences of the United States of America 20080829 36

The phosphodiesterases (PDEs) are metal ion-dependent enzymes that regulate cellular signaling by metabolic inactivation of the ubiquitous second messengers cAMP and cGMP. In this role, the PDEs are involved in many biological and metabolic processes and are proven targets of successful drugs for the treatments of a wide range of diseases. However, because of the rapidity of the hydrolysis reaction, an experimental knowledge of the enzymatic mechanisms of the PDEs at the atomic level is still la ...[more]

PMID: 18757755

Similar Datasets

Structural basis for substrate binding and catalytic mechanism of a human RNA:m5C methyltransferase NSun6.

Project description:5-methylcytosine (m5C) modifications of RNA are ubiquitous in nature and play important roles in many biological processes such as protein translational regulation, RNA processing and stress response. Aberrant expressions of RNA:m5C methyltransferases are closely associated with various human diseases including cancers. However, no structural information for RNA-bound RNA:m5C methyltransferase was available until now, hindering elucidation of the catalytic mechanism behind RNA:m5C methylation. Here, we have solved the structures of NSun6, a human tRNA:m5C methyltransferase, in the apo form and in complex with a full-length tRNA substrate. These structures show a non-canonical conformation of the bound tRNA, rendering the base moiety of the target cytosine accessible to the enzyme for methylation. Further biochemical assays reveal the critical, but distinct, roles of two conserved cysteine residues for the RNA:m5C methylation. Collectively, for the first time, we have solved the complex structure of a RNA:m5C methyltransferase and addressed the catalytic mechanism of the RNA:m5C methyltransferase family, which may allow for structure-based drug design toward RNA:m5C methyltransferase-related diseases.

| S-EPMC5499824 | biostudies-literature

Structural basis of catalytic activation in human splicing.

Project description:Pre-mRNA splicing follows a pathway driven by ATP-dependent RNA helicases. A crucial event of the splicing pathway is the catalytic activation, which takes place at the transition between the activated Bact and the branching-competent B* spliceosomes. Catalytic activation occurs through an ATP-dependent remodelling mediated by the helicase PRP2 (also known as DHX16)1-3. However, because PRP2 is observed only at the periphery of spliceosomes3-5, its function has remained elusive. Here we show that catalytic activation occurs in two ATP-dependent stages driven by two helicases: PRP2 and Aquarius. The role of Aquarius in splicing has been enigmatic6,7. Here the inactivation of Aquarius leads to the stalling of a spliceosome intermediate-the BAQR complex-found halfway through the catalytic activation process. The cryogenic electron microscopy structure of BAQR reveals how PRP2 and Aquarius remodel Bact and BAQR, respectively. Notably, PRP2 translocates along the intron while it strips away the RES complex, opens the SF3B1 clamp and unfastens the branch helix. Translocation terminates six nucleotides downstream of the branch site through an assembly of PPIL4, SKIP and the amino-terminal domain of PRP2. Finally, Aquarius enables the dissociation of PRP2, plus the SF3A and SF3B complexes, which promotes the relocation of the branch duplex for catalysis. This work elucidates catalytic activation in human splicing, reveals how a DEAH helicase operates and provides a paradigm for how helicases can coordinate their activities.

| S-EPMC10208982 | biostudies-literature

Structural basis of peroxidase catalytic cycle of human Prdx6.

Project description:Peroxiredoxin 6 (Prdx6) is a ubiquitously expressed antioxidant non-selenium glutathione peroxidase that is known to play a major role in various physiological and pathological processes. It belongs to the family of peroxidases (referred to as Peroxiredoxins, Prdx's) that work independently of any prosthetic groups or co-factors, and instead utilize a peroxidatic thiol residue for peroxide reduction. Mammalian Prdx's are classified according to the number of Cys implicated in their catalytic activity by the formation of either inter-molecular (typical 2-Cys, Prdx1-4) or intra-molecular (atypical 2-Cys, Prdx5) disulfide bond, or non-covalent interactions (1-Cys, Prdx6). The typical and atypical 2-Prdx's have been identified to show decamer/dimer and monomer/dimer transition, respectively, upon oxidation of their peroxidatic cysteine. However, the alterations in the oligomeric status of Prdx6 as a function of peroxidatic thiol's redox state are still ambiguous. While the crystal structure of recombinant human Prdx6 is resolved as a dimer, the solution structures are reported to have both monomers and dimers. In the present study, we have employed several spectroscopic and electrophoretic probes to discern the impact of change in the redox status of peroxidatic cysteine on conformation and oligomeric status of Prdx6. Our study indicates Prdx6's peroxidase activity to be a redox-based conformation driven process which essentially involves monomer-dimer transition.

| S-EPMC7566464 | biostudies-literature

Structural basis for the catalytic mechanism of mammalian 25-kDa thiamine triphosphatase.

Project description:Mammalian soluble thiamine triphosphatase (ThTPase) is a 25-kDa cytosolic enzyme that specifically catalyzes the conversion of thiamine triphosphate (ThTP) to thiamine diphosphate and has an absolute requirement for divalent cations. We have investigated the kinetic properties of recombinant mouse thiamine triphosphatase (mThTPase) and determined its solution structure by NMR spectroscopy. Residues responsible for binding Mg(2+) and ThTP were determined from NMR titration experiments. The binding of Mg(2+) induced only a minor local conformational change, whereas ThTP binding was found to cause a more global conformational change. We derived a structural model for the mThTPase.ThTP.Mg(2+) ternary complex and concluded from this that whereas free mThTPase has an open cleft fold, the enzyme in the ternary complex adopts a tunnel fold. Our results provide a functional rationale for a number of conserved residues and suggest an essential role for Mg(2+) in catalysis. We propose a mechanism underlying the high substrate specificity of mThTPase and discuss the possible role of water molecules in enzymatic catalysis.

| S-EPMC2447667 | biostudies-literature

Structural basis for a distinct catalytic mechanism in Trypanosoma brucei tryparedoxin peroxidase.

Project description:Trypanosoma brucei, the causative agent of African sleeping sickness, encodes three cysteine homologues (Px I-III) of classical selenocysteine-containing glutathione peroxidases. The enzymes obtain their reducing equivalents from the unique trypanothione (bis(glutathionyl)spermidine)/tryparedoxin system. During catalysis, these tryparedoxin peroxidases cycle between an oxidized form with an intramolecular disulfide bond between Cys(47) and Cys(95) and the reduced peroxidase with both residues in the thiol state. Here we report on the three-dimensional structures of oxidized T. brucei Px III at 1.4A resolution obtained by x-ray crystallography and of both the oxidized and the reduced protein determined by NMR spectroscopy. Px III is a monomeric protein unlike the homologous poplar thioredoxin peroxidase (TxP). The structures of oxidized and reduced Px III are essentially identical in contrast to what was recently found for TxP. In Px III, Cys(47), Gln(82), and Trp(137) do not form the catalytic triad observed in the selenoenzymes, and related proteins and the latter two residues are unaffected by the redox state of the protein. The mutational analysis of three conserved lysine residues in the vicinity of the catalytic cysteines revealed that exchange of Lys(107) against glutamate abrogates the reduction of hydrogen peroxide, whereas Lys(97) and Lys(99) play a crucial role in the interaction with tryparedoxin.

| S-EPMC2662087 | biostudies-literature

Structural basis for DNA 3'-end processing by human tyrosyl-DNA phosphodiesterase 1.

Project description:Tyrosyl-DNA phosphodiesterase (Tdp1) is a DNA 3'-end processing enzyme that repairs topoisomerase 1B-induced DNA damage. We use a new tool combining site-specific DNA-protein cross-linking with mass spectrometry to identify Tdp1 interactions with DNA. A conserved phenylalanine (F259) of Tdp1, required for efficient DNA processing in biochemical assays, cross-links to defined positions in DNA substrates. Crystal structures of Tdp1-DNA complexes capture the DNA repair machinery after 3'-end cleavage; these reveal how Tdp1 coordinates the 3'-phosphorylated product of nucleosidase activity and accommodates duplex DNA. A hydrophobic wedge splits the DNA ends, directing the scissile strand through a channel towards the active site. The F259 side-chain stacks against the -3 base pair, delimiting the junction of duplexed and melted DNA, and fixes the scissile strand in the channel. Our results explain why Tdp1 cleavage is non-processive and provide a molecular basis for DNA 3'-end processing by Tdp1.

| S-EPMC5750209 | biostudies-literature

Structural basis for the catalytic mechanism and α-ketoglutarate cooperativity of glutamate dehydrogenase.

Project description:Glutamate dehydrogenase (GDH) is a key enzyme connecting carbon and nitrogen metabolism in all living organisms. Despite extensive studies on GDHs from both prokaryotic and eukaryotic organisms in the last 40 years, the structural basis of the catalytic features of this enzyme remains incomplete. This study reports the structural basis of the GDH catalytic mechanism and allosteric behavior. We determined the first high-resolution crystal structures of glutamate dehydrogenase from the fungus Aspergillus niger (AnGDH), a unique NADP+-dependent allosteric enzyme that is forward-inhibited by the formation of mixed disulfide. We determined the structures of the active enzyme in its apo form and in binary/ternary complexes with bound substrate (α-ketoglutarate), inhibitor (isophthalate), coenzyme (NADPH), or two reaction intermediates (α-iminoglutarate and 2-amino-2-hydroxyglutarate). The structure of the forward-inhibited enzyme (fiAnGDH) was also determined. The hexameric AnGDH had three open subunits at one side and three partially closed protomers at the other, a configuration not previously reported. The AnGDH hexamers having subunits with different conformations indicated that its α-ketoglutarate-dependent homotropic cooperativity follows the Monod-Wyman-Changeux (MWC) model. Moreover, the position of the water attached to Asp-154 and Gly-153 defined the previously unresolved ammonium ion-binding pocket, and the binding site for the 2'-phosphate group of the coenzyme was also better defined by our structural data. Additional structural and mutagenesis experiments identified the residues essential for coenzyme recognition. This study reveals the structural features responsible for positioning α-ketoglutarate, NADPH, ammonium ion, and the reaction intermediates in the GDH active site.

| S-EPMC5925808 | biostudies-literature

Structural basis and catalytic mechanism for the dual functional endo-beta-N-acetylglucosaminidase A.

Project description:Endo-beta-N-acetylglucosaminidases (ENGases) are dual specificity enzymes with an ability to catalyze hydrolysis and transglycosylation reactions. Recently, these enzymes have become the focus of intense research because of their potential for synthesis of glycopeptides. We have determined the 3D structures of an ENGase from Arthrobacter protophormiae (Endo-A) in 3 forms, one in native form, one in complex with Man(3)GlcNAc-thiazoline and another in complex with GlcNAc-Asn. The carbohydrate moiety sits above the TIM-barrel in a cleft region surrounded by aromatic residues. The conserved essential catalytic residues - E173, N171 and Y205 are within hydrogen bonding distance of the substrate. W216 and W244 regulate access to the active site during transglycosylation by serving as "gate-keepers". Interestingly, Y299F mutation resulted in a 3 fold increase in the transglycosylation activity. The structure provides insights into the catalytic mechanism of GH85 family of glycoside hydrolases at molecular level and could assist rational engineering of ENGases.

| S-EPMC2646837 | biostudies-literature

Molecular basis for the catalytic mechanism of human neutral sphingomyelinases 1 (hSMPD2).

Project description:Enzymatic breakdown of sphingomyelin by sphingomyelinase (SMase) is the main source of the membrane lipids, ceramides, which are involved in many cellular physiological processes. However, the full-length structure of human neutral SMase has not been resolved; therefore, its catalytic mechanism remains unknown. Here, we resolve the structure of human full-length neutral SMase, sphingomyelinase 1 (SMPD2), which reveals that C-terminal transmembrane helices contribute to dimeric architecture of hSMPD2 and that D111 - K116 loop domain is essential for substrate hydrolysis. Coupled with molecular docking, we clarify the binding pose of sphingomyelin, and site-directed mutagenesis further confirms key residues responsible for sphingomyelin binding. Hybrid quantum mechanics/molecular mechanics (QM/MM) molecular dynamic (MD) simulations are utilized to elaborate the catalysis of hSMPD2 with the reported in vitro substrates, sphingomyelin and lyso-platelet activating fator (lyso-PAF). Our study provides mechanistic details that enhance our knowledge of lipid metabolism and may lead to an improved understanding of ceramide in disease and in cancer treatment.

| S-EPMC10682184 | biostudies-literature

Structural basis for catalytic activation by the human ZNF451 SUMO E3 ligase.

Project description:E3 protein ligases enhance transfer of ubiquitin-like (Ubl) proteins from E2 conjugating enzymes to substrates by stabilizing the thioester-charged E2~Ubl in a closed configuration optimally aligned for nucleophilic attack. Here, we report biochemical and structural data that define the N-terminal domain of the Homo sapiens ZNF451 as the catalytic module for SUMO E3 ligase activity. The ZNF451 catalytic module contains tandem SUMO-interaction motifs (SIMs) bridged by a Pro-Leu-Arg-Pro (PLRP) motif. The first SIM and PLRP motif engage thioester-charged E2~SUMO while the next SIM binds a second molecule of SUMO bound to the back side of E2. We show that ZNF451 is SUMO2 specific and that SUMO modification of ZNF451 may contribute to activity by providing a second molecule of SUMO that interacts with E2. Our results are consistent with ZNF451 functioning as a bona fide SUMO E3 ligase.

| S-EPMC4709122 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data