Результаты поиска

B56δ long-disordered arms form a dynamic PP2A regulation interface coupled with global allostery and Jordan's syndrome mutations.

Intrinsically disordered regions (IDR) and short linear motifs (SLiMs) play pivotal roles in the intricate signaling networks governed by phosphatases and kinases. B56δ (encoded by PPP2R5D) is a regulatory subunit of protein phosphatase 2A (PP2A) with long IDRs that harbor a substrate-mimicking SLiM and multiple phosphorylation sites. De novo missense mutations in PPP2R5D cause intellectual disabilities (ID), macrocephaly, Parkinsonism, and a broad range of neurological symptoms. Our single-particle cryo-EM structures of the PP2A-B56δ holoenzyme reveal that the long, disordered arms at the B56

Effect of treatment with pyridoxine on aspartate aminotransferase activities in pyridoxine-deficient rat tissues.

In rat liver, 90% of the aspartate aminotransferase is present as the holoenzyme. In pyridoxine deficiency, the ratio of holoenzyme activity to total activity is markedly reduced, but after pyridoxine injection it was found to rapidly increase, although the total enzyme activity remained low for a few days. The activities of aspartate aminotransferase isozymes in pyridoxine-deficient rat tissues and the effect of pyridoxine treatment on their activities were examined. The intestinal enzyme activities of pyridoxine-deficient rats were readily reconstituted in the presence of pyridoxal phosphate

N3A motifs in RIβ mediate allosteric crosstalk between cAMP and ATP in PKA activation.

The RIβ subunit of cAMP-dependent protein kinase (PKA) is highly expressed in the brain, yet it remains the least studied of the PKA regulat … inker and show how each contributes in a unique but coordinated way to allosteric activation of RIβ holoenzymes by cAMP. A key difference in our RIβ:C structure is the rotation of the side chain of W260 at the N-terminus of the αA Helix in N3AB. W260, at the R:C interface in the holoenzyme, is also the capping residue for cAMP bound to CNB-A, so we may have a …

Crystallization and preliminary X-ray diffraction analysis of the hyperthermostable NAD-dependent glutamate dehydrogenase from Pyrobaculum islandicum.

NAD-dependent glutamate dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum was crystallized in the apo- and holoenzyme forms. Crystals were obtained using 2-propanol and polyethylene glycol MME 550 as precipitants for the apoenzyme and holoenzyme, respectively. The apoenzyme crystals belong to the trigonal space group P3(1)21 or its enantiomorph P3(2)21. The asymmetric unit contains three subunits; the values of the Matthews coefficient (VM) and the solvent content are 2.9 A3 Da-1 and 57%, respectively. A native data set was collected to a highest resolution limit of 4.0

[Alpha-ketoglutarate dehydrogenase activity of Candida lipolytica during alpha-ketoglutarate biosynthesis].

The activity of alpha-ketoglutarate dehydrogenase (KGDH) was assayed in cell-free yeast homogenates with respect to studying the mechanism of alpha-ketoglutarate (KG) overproduction by the thiamine-heterotrophic culture of Candida lipolytica in media with hexadecane. Two types of yeast cells were used for analysis: (1) cells grown in the conditions of thiamine excess (500 mcg/l) and taken at the logariphmic growth phase; (2) cells grown in the conditions of thiamine deficiency and taken at the beginning of KG overproduction when the cultural growth was limited by thiamine. The activity of KGDH

Binding of transcriptional activators to sigma 54 in the presence of the transition state analog ADP-aluminum fluoride: insights into activator mechanochemical action.

Conformational changes in sigma 54 (sigma(54)) and sigma(54)-holoenzyme depend on nucleotide hydrolysis by an activator. We now show that sigma(54) and its holoenzyme bind to the central ATP-hydrolyzing domains of the transcriptional activators PspF and NifA in the presence of ADP-aluminum fluoride, an analog of ATP in the transition state for hydrolysis. Direct binding of sigma(54) Region I to activator in the presence of ADP-aluminum fluoride was shown and inferred from in vivo suppression genetics. Energy transduction appears to occur through activator contacts to sigma(54) Region I. ADP-al

Crystallization and preliminary X-ray analysis of (1R,6R)-2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC) synthase (MenD) from Escherichia coli.

(1R,6R)-2-Succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC) synthase, also called MenD, participates in the menaquinone (vitamin K2) biosynthetic pathway. The enzyme is a part of the superfamily of ThDP-dependent enzymes; however, it is the only enzyme known to catalyze a Stetter-like 1,4-addition of a ThDP adduct to the beta-carbon of an unsaturated carboxylate. This is the first reported crystallization of the apoenzyme and holoenzyme forms of MenD. The apoenzyme crystals were obtained by sitting-drop vapour diffusion with 70% MPD. However, the crystals were too small to collect di

Inorganic carbon levels regulate growth via SigC signaling cascade in cyanobacteria.

Cyanobacterial growth depends on inorganic carbon (Ci; CO2 and bicarbonate) concentration, but mechanism(s) adjusting photosynthesis and growth according to Ci remain unclear. ΔrpoZ cells lacking the ω subunit of the RNA polymerase (RNAP) show a unique high-CO2 lethal phenotype in Synechocystis sp. PCC 6803. Bioinformatics, biochemical and 3D modeling studies were used to reveal how suppressor mutations rescue ΔrpoZ cells in 3% CO2. Suppressor mutations were mapped to the ssr1600 gene. Ssr1600 was shown to function as an anti-σ factor antagonist. The Slr1861 protein was identified as an anti-σ

Inhibition of delta-aminolevulinate dehydratase in trichloroethylene-exposed rats, and the effects on heme regulation.

A pronounced and irreversible depression of the erythroid and liver delta-aminolevulinate dehydratase (porphobilinogen synthase; 5-aminolevulinate hydro-lyase, EC 4.2.1.24) activity was observed in rats exposed to trichloroethylene, a widely used solvent. The depression could not be restored after the treatment with dithiothreitol and zinc; however, radioimmunoassay of delta-aminolevulinate dehydratase indicated that trichloroethylene exposure did not essentially decrease the amount of enzyme. The depression of the enzyme activity thus proved to be due not to a reduction in the enzyme amount b

Interactions between large and small subunits of different acetohydroxyacid synthase isozymes of Escherichia coli.

The large, catalytic subunits (LSUs; ilvB, ilvG and ilvI, respectively) of enterobacterial acetohydroxyacid synthases isozymes (AHAS I, II a … only approximately 15, <<1 and approximately 3%, respectively, of the activity of their respective holoenzymes, but the holoenzymes can be reconstituted with complete recovery of activity. We have examined the activation of each of the LSUs by SSUs from different isozymes and ask to what extent such activation is specific; that is, is effective nonspecific i …

Biosynthesis of casein kinase II in lymphoid cell lines.

We have analyzed the biosynthesis of casein kinase II. In exponentially growing tissue culture cells, the beta subunit was synthesized in excess of the catalytic subunit (alpha). A substantial fraction of newly synthesized beta was degraded within the first hour. The remaining fraction of beta was incorporated into holoenzyme. In contrast, little degradation of newly synthesized alpha subunit was observed and most was quickly and efficiently incorporated into holoenzyme. The assembly of beta with alpha was paralleled by an increase in apparent molecular mass of beta due to phosphorylation. The

Подъемные элементы редукции Ribunocleotide из желтой лихорадки комаров, Aedes aegypti: клонирование и выражение.

Редуктаза Рибонуклеотида стимулирует деново синтез деоксирибонуклеотидов. Редукции первого класса используют железный центр для создания тиросилового свободного радикала, который может инициировать образование дезоксирибонуклеотида. Эти редукции представляют собой альфа-2 бета-2 голоэнзимы, а субъекты обозначаются как R1 и R2. R1 содержит аллостерический связной участок и активный объект, в то время как R2 содержит биядерный железный центр, который инициирует образование тиросил радикал. Мы клонировали и секвенировали CDNA, кодируя субъект R1 и R2 в желтой лихорадке комаров, Aedes aegypti. Свя

Совпадение в аннотации оригинала
Ribonucleotide reductase catalyses the de novo synthesis of deoxyribonucleotides. Class I reductases use an iron center to generate a tyrosyl free radical that can initiate formation of the deoxyribonucleotide. These reductases are alpha 2 beta 2 holoenzymes, and the subunits are denoted as R1 and R2. R1 contains the allosteric binding site and the active si…

Regulation between LRRK2 and PP2A signaling in cellular models of Parkinson's disease.

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most frequent cause of late-onset familial and idiopathic Parkinson's disease (PD), known to date. Importantly, recent data from postmortem tissue as well as biomarker studies suggest that independent of mutations, increased kinase activity of LRRK2 plays an essential role in idiopathic PD pathogenesis. Despite extensive research on LRRK2, its activation mechanism(s) and how the various mutations result in increased kinase activity and neuronal death are still not completely understood. Accumulating evidence points to LRRK2 phosphoregul

The modification of DNA-dependent RNA polymerase from Escherichia coli by an alkylating derivative of rifamycin SV.

3-(2-Bromo[1-14C]acetamidoethyl)-thio-rifamycin SV, abbreviated BrAcNEtS-Rif, and alkylating derivative of rifamycin SV was synthesized. A four-fold excess of BrAcNEtS-Rif inhibited the enzymic activity of RNA polymerase from Escherichia coli to 97%. Incubation of RNA polymerase with Br[14C]AcNEtS-Rif led to covalent substitution. The reaction of Br[14C]AcNEtS-Rif with enzyme at a ratio of 1.4:1 and a concentration of 63 nM was found to proceed with a half life of 1 h at 37 degrees C. The enzyme could be protected from reaction with BrAcNEtS-Rif by either rifampicin or the hybrid [poly(dT)]-[r

Acrodysostosis type 1: mechanisms explaining PRKAR1A mutation mediated dysregulation of cAMP-PKA signalling.

Acrodysostosis type 1 (ACRDYS1) is a rare multisystem developmental disorder affecting skeletal growth, endocrine function, neurodevelopment … nits. Importantly, mutant RIα is expressed at near-normal levels and assembles efficiently into PKA holoenzymes, but these complexes respond weakly and sluggishly to physiological cAMP signals. Drawing on structural, biochemical, cellular, and in vivo studies, we define a dual pathogenic mechanism underlying ACRDYS1. First, defective cAMP-driven conformationa …

Activators of phosphorylase kinase alter the cross-linking of its catalytic subunit to the C-terminal one-sixth of its regulatory alpha subunit.

Phosphorylase kinase, a regulatory enzyme of glycogenolysis in skeletal muscle, is a hexadecameric oligomer consisting of four copies each of a catalytic subunit (gamma) and three regulatory subunits (alpha, beta, and delta, the last being endogenous calmodulin). The enzyme is activated by a variety of effectors acting through its regulatory subunits. To probe the quaternary structure of nonactivated and activated forms of the kinase, we used the heterobifunctional, photoreactive cross-linker N-5-azido-2-nitrobenzoyloxysuccinimide. Mono-derivatization of the holoenzyme with the succinimidyl gr

Selective Inhibition of CK2: Emerging Strategies and Future Directions.

CK2 is a constitutively active serine/threonine kinase implicated in cancer, viral infection, and neurodegeneration, but its conserved ATP-binding site and complex holoenzyme assembly have long made selective inhibition challenging. This Perspective discusses three complementary strategies for selective CK2 inhibition: targeting the CK2α/β interface with small molecules, disrupting the same interface with conformationally constrained peptides, and exploiting the cryptic αD pocket for dual-site inhibitor design. CK2α/β-interface inhibitors, including CAM187, CAM7117 and P8C9, established struct

Allosteric modulation of the RNA polymerase catalytic reaction is an essential component of transcription control by rifamycins.

Rifamycins, the clinically important antibiotics, target bacterial RNA polymerase (RNAP). A proposed mechanism in which rifamycins sterically block the extension of nascent RNA beyond three nucleotides does not alone explain why certain RNAP mutations confer resistance to some but not other rifamycins. Here we show that unlike rifampicin and rifapentin, and contradictory to the steric model, rifabutin inhibits formation of the first and second phosphodiester bonds. We report 2.5 A resolution structures of rifabutin and rifapentin complexed with the Thermus thermophilus RNAP holoenzyme. The str

Retracted: Allosteric Activators of Protein Phosphatase 2A Display Broad Antitumor Activity Mediated by Dephosphorylation of MYBL2.

Protein phosphatase 2A (PP2A) enzymes can suppress tumors, but they are often inactivated in human cancers overexpressing inhibitory protein … ther cancer cells in prometaphase. In contrast, SMAPs, a separate class of compounds, activate PP2A holoenzymes containing a different regulatory subunit, do not dephosphorylate MYBL2, and arrest tumor cells in G1 phase. Our findings demonstrate that small molecules can serve as allosteric switches to activate distinct PP2A complexes with unique substrates.

The Multifaceted Role of Protein Phosphatase 1 in Plasmodium.

Protein phosphatase type 1 (PP1) forms a wide range of Ser/Thr-specific phosphatase holoenzymes which contain one catalytic subunit (PP1c), present in all eukaryotic cells, associated with variable subunits known as regulatory proteins. It has recently been shown that regulators take a leading role in the organization and the control of PP1 functions. Many studies have addressed the role of these regulators in diverse organisms, including humans, and investigated their link to diseases. In this review we summarize recent advances on the role of PP1c in Plasmodium, its interactome and regulator…