SEARCH · Search PubMed

Results for “Enzyme Reactivators”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 343 records · Page 19Linked to original sources

PubMed1992

Characterization of mutant forms of the quinoprotein methanol dehydrogenase lacking an essential calcium ion.

Methanol dehydrogenase (MDH) from Methylobacterium extorquens, Methylophilus methylotrophus, Paracoccus denitrificans and Hyphomicrobium X all contained a single atom of Ca2+ per alpha 2 beta 2 tetramer. The role of Ca2+ was investigated using the MDH from Methylobacterium extorquens. This was shown to be similar to the MDH from Hyphomicrobium X in having 2 mol of prosthetic group (pyrroloquinoline quinine; PQQ) per mol of tetramer, the PQQ being predominantly in the semiquinone form. MDH isolated from the methanol oxidation mutants MoxA-, K- and L- contained no Ca2+. They were identical with the enzyme isolated from wild-type bacteria with respect to molecular size, subunit configuration, pI, N-terminal amino acid sequence and stability under denaturing conditions (low pH, high urea and high guanidinium chloride) and in the nature and content of the prosthetic group (2 mol of PQQ per mol of MDH). They differed in their lack of Ca2+, the oxidation state of the extracted PQQ (fully oxidized), absence of the semiquinone form of PQQ in the enzyme, reactivity with the suicide inhibitor cyclopropanol and absorption spectrum, which indicated that PQQ is bound differently from that in normal MDH. Incubation of MDH from the mutants in calcium salts led to irreversible time-dependent reconstitution of full activity concomitant with restoration of a spectrum corresponding to that of fully reduced normal MDH. It is concluded that Ca2+ in MDH is directly or indirectly involved in binding PQQ in the active site. The MoxA, K and L proteins may be involved in maintaining a high Ca2+ concentration in the periplasm. It is more likely, however, that they fill a 'chaperone' function, stabilizing a configuration of MDH which permits incorporation of low concentrations of Ca2+ into the protein.

Alcohol Oxidoreductases↗

PubMed1978

Is high-molecular-weight-renin binding of renin to the protease inhibitors and lipoproteins?

1. Two high-molecular-weight forms of renin (molecular weights 800 000 and 70 000) are present in mouse plasma. 2. The 800 000 form could be activated and converted into the fully active 40 000 form, by acid or limited proteolysis. The 70 000 form was activated without change in molecular weight. 3. In addition to its enzymic activity, renin was measured by a direct radioimmunoassay, which revealed that the current acid treatment of plasma did not activate all the renin present. 4. Renin is stored as fully active 40 000 renin, with a specific enzymic reactivity of 0.4 times 10(-3) GU ng(-1), in the submaxillary gland of mice. 5. Pure 125I-labelled 40 000 submaxillary renin did not bind to plasma proteins. However, by changing the tertiary structure of renin, it was bound to some of the plasma protease inhibitors; alpha2-macroglobulin, inter-alpha-trypsin inhibitor and alpha2-antithrombin. It was also bound to alpha1- and beta1-lipoprotein, albumin and an unidentified plasma protein. No binding was seen to more than 50 other studied plasma proteins.

Animals↗

PubMed2002

Assessment of factor V, VII and X activities, the key coagulant proteins of the tissue factor pathway in poultry plasma.

1. Assay methods were developed for key components of the tissue factor pathway of blood coagulation, namely Factor V, Factor VII and Factor X. Using these assays, plasma from healthy laying hens, cockerels and broilers was shown to contain functional and equivalent amounts of each of these clotting factors. 2. The plasma activities for Factor V, Factor VII and Factor X can only be accurately determined when chicken tissue factor is used to initiate the coagulation mechanism in poultry plasma. Neither human tissue factor nor rabbit tissue factor forms a fully functional enzyme reactive complex with chicken Factor VII. 3. The overall tissue factor pathway coagulation mechanism was evaluated in plasma from laying hens, cockerels and broilers using the one-stage prothrombin time assay. As long as sufficient tissue factor was used, the overall clotting time results obtained with human recombinant tissue factor were not significantly different from those obtained with chicken tissue factor. 4. We conclude that poultry plasma does possess a fully functional tissue factor coagulation mechanism, but homologous chicken tissue factor must be used for in vitro assays of the components of this pathway.

Animals↗

PubMed2007

IFNgamma: issuing macrophages a license to kill.

T cells tell macrophages when to start making the toxic soup of lysosomal enzymes, reactive oxygen species, and nitric oxide that destroys intracellular pathogens. In 1983, Carl Nathan proved that this start signal comes in the form of the secreted cytokine IFNgamma.

Animals↗

PubMed1975

Isolation and study of mutants lacking a derepressible phosphatase in Chlamydomonas reinhardi.

In the green alga Chlamydomonas reinhardi, removal of inorganic phosphate from the culture medium results in the increase of phosphatase activity (derepression) in the wild-type (WT) strain as well as in a double mutant (P2Pa)) lacking the two main constitutive acid phosphatases. Following treatment of WT and P2Pa with N-methyl-N-nitro-N-nitrosoguanidine (MNNG), mutants were recovered which display very low phosphatase activities when grown in the absence of phosphate; as shown by electrophoresis, they lack one non-migrating phosphatase (PD mutants). This enzyme is active over a wide range of pH with an optimum at pH 7.5. The comparison of elctropherograms form WT and mutants grown on media with or without phosphate allowed us to provide a tentative definition of the pool of derepressible phosphatases in Chlamydomonas: in addition tothe neutral phosphatase lacking in PD mutants, Chlamydomonas produces two electrophoretic forms of alkaline phosphatase showing an optimal activity at pH 9.5.

Acid Phosphatase↗

PubMed1987

Properties of DNase I digestion of the deoxyoligonucleotide: 5'd(ATCGTACGAT)2(3').

Deoxyribonuclease I digestion of the deoxyoligodecamer 5'd(ATCGTACGAT)2(3') has been examined in detail to study the kinetic and structural properties of this enzyme substrate system in solution. In addition, these studies have defined, in general, those DNase I conditions to be used in future drug-DNA footprinting experiments. Special attention has been taken of those properties of DNase I that are critical for quantitation of ligand binding to small DNA fragments, and that aid in designing oligomers to be used in footprinting experiments. Enzyme activity was observed at all phosphodiester bonds in the decamer studied with varying affinity, except for the first four bonds at the 5' end of the oligomer. The DNA substrate concentration is always in excess, in order to achieve conditions of no more than one DNase I cleavage per DNA molecule. Reactions were controlled so that 65% or more of the initial amount of decamer substrate remained after DNase I digestion. It was observed that the rate of enzyme reactivity decreases with digestion time and is sensitive to the experimental conditions.

Autoradiography↗

PubMed2002

Intravenous L-lactate application in minipigs partially protects acetylcholinesteratic but not butyrylcholinesteratic activity in plasma from inhibition by paraoxon.

OBJECTIVE: Intoxications with organophosphorous compounds such as paraoxon, an inhibitor of serine hydrolases, mainly butyrylcholinesterase and acetylcholinesterase, are frequent. Oximes are the only enzyme reactivators clinically available. In vitro studies have shown that L(+)-lactate reduces the inhibition of acetylcholinesteratic (AChEA) and butyrylcholinesteratic activity of plasma (BChEA) by paraoxon. DESIGN: The purpose of this in vivo study was to determine whether intravenous L(+)-lactate application under normoxic/normocapnic/normohydrogenemic conditions is able to protect AChEA and BChEA from paraoxon inhibition. SETTING: University research institute. SUBJECTS: Eighteen female minipigs. INTERVENTIONS: Animals were anesthetized, intubated, and mechanically ventilated. Every animal received 1 mg of paraoxon per kilogram of body weight in 50 mL of saline over 50 mins. In addition to receiving paraoxon, six pigs of 18 received 2.5 g (0.125 g kg-1 of body weight) of intravenous L(+)-lactate in 50 mL of saline over 50 mins, and six other pigs received 10 g of L(+)-lactate (0.5 g kg-1 of body weight), whereas the six remaining served as controls. MEASUREMENTS AND MAIN RESULTS: In central venous blood, plasma acetylcholinesteratic and butyrylcholinesteratic activity were measured before paraoxon (baseline, 0 mins), immediately after paraoxon (50 mins after start), and 110, 170, 230, 290, 530, and 1010 mins after the start of infusion. Although 10 g of intravenous L(+)-lactate application had a statistically significant protective effect in vivo on AChEA, 2.5 g did not. No significant protective effect on BChEA was achieved with either 2.5 g or 10 g of L(+)-lactate. CONCLUSIONS: Ten grams of L(+)-lactate can increase AChEA when administered simultaneously with paraoxon. Further study of the in vivo effects of L(+)-lactate after paraoxon intoxication and a formal comparison with standard oxime therapy seem warranted. Also, methods for achieving a prolonged elevated lactate concentration in vivo should be investigated.

Acetylcholinesterase↗

PubMed1983

Neuronophagia by leukocytes in experimental spinal cord injury.

Experimental spinal cord injury was produced in cats by compression trauma. The major histologic features in the first 24 hours following injury included hemorrhagic necrosis, edema, and acute inflammation of gray and white matter. Neutrophilic leukocytes (NL) were apparent in the walls of and adjacent to veins and venules within four hours of injury, but large numbers of NL were not observed in tissue until eight to 24 hours. Acute inflammation was especially prominent in gray matter, and NL frequently surrounded and phagocytized neuronal somata. Large numbers of NL and neuronophagia by leukocytes were evident only in areas of hemorrhage. The role of blood in producing the acute inflammatory response and in generating chemotactic factors responsible for neuronophagia is incompletely understood. Neutrophilic leukocytes, in addition to their response as phagocytes, release histolytic enzymes, reactive species of oxygen, and proinflammatory factors which lead to further tissue necrosis and inflammation.

Animals↗

PubMed1995

Signaling mechanisms in human neutrophils.

Leukocytes possess many properties critical to their effective functioning in inflammation, including the ability to migrate to the site of inflammation and release an impressive armamentarium of toxic products such as proteolytic enzymes, reactive oxygen species, and cationic proteins, capable of killing invading pathogens. This review focuses on the transmembrane signaling events whereby factors present in an inflammatory milieu activate these leukocyte effector functions. In the past several years, many of the components of these pathways have been elucidated at the molecular level, but large gaps remain in our understanding. The discussion follows the path beginning from the exofacial side of the plasma membrane toward the cell interior: from membrane receptors, GTP-binding proteins and adapter proteins to intermediary pathways including phospholipases and protein kinases. A detailed understanding of these regulatory mechanisms will have important therapeutic implications for amelioration of inflammatory tissue injury.

Animals↗

PubMed2005

Telomerase and cancer stem cells.

Telomerase is critical for the integrity of stem cell compartments. Mutations in telomerase components lead to telomere shortening and hematopoietic stem cell failure in autosomal dominant dyskeratosis congenita and aplastic anemia. Telomerase activity is readily detected in most cancers but not in adult somatic cells. The telomere hypothesis for cancer states that telomerase is reactivated in late stages of carcinogenesis. However, recent evidence has suggested a stem cell origin for certain cancers, implying that the genetic alterations that lead to cancer accumulate in tissue-specific stem cells and not in adult somatic cells. In these cancers, stem cells would already have telomerase and it would not need to be reactivated. Here, we reconsider the telomere hypothesis in view of this evidence and propose that, rather than telomerase reactivation, enzyme activity may increase in later stages of carcinogenesis due to increased expression or efficient assembly of telomerase components. Understanding these mechanisms will refine approaches to telomerase inhibition in cancer.

Dyskeratosis Congenita↗

PubMed2026

TUBA4A Pathogenic Variant Manifesting With Adulthood-Onset Genetic Myasthenic Syndrome, Myopathy, and Infertility.

OBJECTIVES: TUBA4A pathogenic variants are associated with ALS, frontotemporal dementia, spastic ataxia, spasticity, ataxia, Parkinson's disease, female infertility, macrothrombocytopenia, and myopathy. Four recently reported patients with TUBA4A neonatal/childhood onset myopathy had also a decrement on repetitive nerve stimulation (RNS), but such a finding was not further characterized. We describe a patient with a TUBA4A pathogenic variant with adulthood-onset genetic myasthenic syndrome accompanied by myopathy and infertility to highlight the neuromuscular junction defect as the main feature of the patient's phenotype. METHODS: We reviewed the patient's clinical and laboratory findings and performed transcriptomic analysis on the patient's muscle. RESULTS: A 53-year-old woman with infertility of unknown etiology manifested fatigability and proximal upper limb muscle weakness in her mid-30s, followed by lower limb involvement. Her examination showed proximal muscle weakness and fatigability but spared facial muscles. CK values were mildly elevated. Anti-AChR, MuSK, P/Q-type calcium channel, and LRP4 antibodies were absent. 2 Hz RNS showed decrement (-14% to -48%) in limb muscles that improved with 3,4-dyaminopyridine (3,4-DAP). Facilitation (231%) occurred in the trapezius. Muscle biopsy showed patchy loss of oxidative enzyme reactivity and no C5b9 or IgG at neuromuscular junctions. Whole genome sequencing identified a heterozygous known TUBA4A pathogenic variant (c.850G>A, p.Glu284Lys). Patient improved with 3,4-DAP and albuterol. DISCUSSION: TUBA4A p.Glu284Lys can lead to treatable myasthenic syndrome with postsynaptic and likely presynaptic involvement, as suggested by the patient's electrophysiological findings and response to therapy. This patient expands the TUBA4A-disorder spectrum to include overlapping myasthenic syndrome-myopathy and shows that neuromuscular disease and infertility can occur within the same patient.

Humans↗

PubMed1993

Multiphasic denaturation of glutathione transferase B1-1 by guanidinium chloride. Role of the dimeric structure on the flexibility of the active site.

The unfolding and refolding mechanisms of dimeric glutathione transferase GSTB1-1 from Proteus mirabilis, using guanidinium chloride as a denaturant, have been investigated. The protein transitions were monitored by enzyme activity, intrinsic fluorescence, far ultraviolet circular dichroism and gel-filtration chromatography. The non coincidence of denaturation curves at equilibrium indicates that the unfolding of GSTB1-1 is a multistep process, i. e. inactivation of the structured dimer, dissociation into partially structured monomers followed by complete unfolding. In the 50% inactivated enzyme the Km for glutathione increases threefold, while the kcat appears almost the same, indicating that the initial phase of the denaturation involves the binding site of glutathione. The rapid recovery of the folded dimer precedes the complete enzyme reactivation. This indicates that the reconstitution of the native structure of GSTB1-1 is the result of folding and association of compact monomers followed by subtle rearrangements of assembled monomers that build up the active site.

Binding Sites↗

PubMed1995

Biochemical characterization of the 8-hydroxy-5-deazaflavin-reactive hydrogenase from Methanosarcina barkeri Fusaro.

The membrane-associated coenzyme F420-reactive hydrogenase of the anaerobic methanogenic archaeon Methanosarcina barkeri Fusaro has been purified 95-fold to apparent homogeneity. A new purification procedure and altered storage conditions gave substantially higher yield (13.4% versus 4.3%) and specific coenzyme F420-reducing activity (82.8 mumol.min-1.mg protein-1 versus 11.5 mumol.min-1.mg protein-1) than reported previously [Fiebig, K. & Friedrich, B. (1989) Eur. J. Biochem. 184, 79-88]. The predominant coenzyme F420-reactive form of the hydrogenase has an apparent molecular mass of 198 kDa and is composed of three non-identical subunits with apparent molecular masses of 48 (alpha), 33 (beta), and 30 kDa (gamma), apparently in a stoichiometry of alpha 2 beta 2 gamma 1. This minimal coenzyme F420-reducing hydrogenase formed aggregates with apparent molecular masses of approximately 845 kDa. 1 mol of the 198-kDa form of hydrogenase contained 2 mol FAD, 2 mol nickel, 28-32 mol non-heme iron, and 34 mol acid-labile sulfur; in addition, 0.2 mol selenium was detected. The isoelectric point was 5.30. The amino acid sequence PXXRXEGH, where X is any amino acid, was found to be conserved in the N-termini of the putative nickel-binding subunits of most [NiFe]- and [NiFeSe]hydrogenases of methanogenic Archaea and Bacteria. However, this motif was not detected in the protein sequences of [Fe]hydrogenases. Maximal coenzyme F420-reducing activity was obtained with reductively reactivated enzyme at 55 degrees C in the pH range 6.5-7.25. The Km values of the purified enzyme for H2 with coenzyme F420 or methylviologen as electron acceptor were extremely low, namely 3 microM and 4 microM. The catalytic efficiency coefficients (kcat/Km) for H2 with both reducible cosubstrates were high: 2.5 x 10(7) M-1.s-1 with coenzyme F420 and 6.9 x 10(7) M-1.s-1 with methylviologen.

Amino Acid Sequence↗

PubMed1994

Stability and activity of hydrogenases of Methanobacterium thermoautotrophicum and Alcaligenes eutrophus in reversed micellar systems.

In water-in-oil microemulsion the membrane-associated F420-hydrogenase of Methanobacterium thermoautotrophicum (strain Marburg) and the membrane-bound hydrogenase of Alcaligenes eutrophus H 16 (MBH) showed prolonged activity at elevated temperatures (measured as hydrogen production) as compared to aqueous buffer solution. The temperature optimum of the reactions was about 15 degrees C higher than in aqueous buffer solution. Activity of the almost completely inactivated F420-hydrogenase could be partially recovered by transfer into microemulsion.

Alcaligenes↗

PubMed1997

Gingival crevicular interleukin-1 and interleukin-1 receptor antagonist levels in periodontally healthy and diseased sites.

Interleukin-1 (IL-1) molecules, IL-1 alpha and IL-1 beta are cytokines involved in the acute-phase response against infection and in the pathogenesis of periodontal destruction. Administration of exogenous IL-1 receptor antagonist (IL-1ra) is effective in reducing the inflammatory reactions mediated by IL-1. However, the relationship between these three naturally occurring IL-1 molecules and periodontal diseases has been poorly characterized. We investigated the correlation of gingival crevicular IL-1 molecules and the clinical status of patients with different severities of periodontitis. IL-1 alpha, IL-1 beta, IL-1ra and the total IL-1/IL-1ra ratio (IL-1 activity index; IL-1AI) were measured in 75 gingival crevicular fluid (GCF) samples from non-inflamed gingiva sites in 2 healthy subjects and diseased sites in 7 patients with several types of periodontitis. IL-1 alpha, IL-1 beta and IL-1ra were measured by specific non-cross-reactive enzyme linked immunosorbent assay. The probing depth, gingival index and alveolar bone loss of each site was recorded at the time of GCF sampling. The total amount of IL-1 alpha, IL-1 beta and the IL-1AI, but not total IL-1ra, were found to be correlated with alveolar bone loss score. Three IL-1 molecules were also measured in the gingival tissue of patients with periodontitis. A similar progressive decrease of the IL-1AI was detected in gingival tissue with periodontitis. These results suggest that the amounts of both crevicular IL-1 and IL-1AI are closely associated with periodontal disease severity.

Acute-Phase Reaction↗

PubMed1990

Anatomy of a conformational change: hinged "lid" motion of the triosephosphate isomerase loop.

Triosephosphate isomerase (TIM) is used as a model system for the study of how a localized conformational change in a protein structure is produced and related to enzyme reactivity. An 11-residue loop region moves more than 7 angstroms and closes over the active site when substrate binds. The loop acts like a "lid" in that it moves rigidly and is attached by two hinges to the remainder of the protein. The nature of the motion appears to be built into the loop by conserved residues; the hinge regions, in contrast, are not conserved. Results of molecular dynamics calculations confirm the structural analysis and suggest a possible ligand-induced mechanism for loop closure.

Amino Acid Sequence↗

PubMed1990

Partial Purification and Characterization of Two Hydrogenases from the Extreme Thermophile Methanococcus jannaschii.

F(420)-nonreactive and F(420)-reactive hydrogenases have been partially purified from Methanococcus jannaschii, an extremely thermophilic methanogen isolated from a submarine hydrothermal vent. The molecular weights of both hydrogenases were determined by native gradient electrophoresis in 5 to 27% polyacrylamide gels. The F(420)-nonreactive hydrogenase produced one major band (475 kilodaltons), whereas the F(420)-reactive hydrogenase produced two major bands (990 and 115 kilodaltons). The F(420)-nonreactive hydrogenase consisted of two subunits (43 and 31 kilodaltons), and the F(420)-reactive hydrogenase contained three subunits (48, 32, and 25 kilodaltons). Each hydrogenase was active at very high temperatures. Methyl viologen-reducing activity of the F(420)-nonreactive hydrogenase was maximal at 80 degrees C but was still detectable at 103 degrees C. The maximum activities of F(420)-reactive hydrogenase for F(420) and methyl viologen were measured at 80 and 90 degrees C, respectively. Low but measureable activity toward methyl viologen was repeatedly observed at 103 degrees C. Moreover, the half-life of the F(420)-nonreactive hydrogenase at 70 degrees C was over 9 h, and that of the F(420)-reactive enzyme was over 3 h.

Journal Article↗

PubMed1967

Distribution of cytochrome c peroxidase activity in wild-type and petite cells of bakers' yeast grown aerobically and anaerobically.

Studies of mitochondrial biogenesis in yeast have been hampered by a lack of suitable membrane markers in anaerobically grown cells subsequently grown in air. Cytochrome c peroxidase activity and subcellular location was studied to determine whether it would be a useful marker for an analysis of mitochondrial formation. Cytochemical tests revealed enzyme reaction product on all mitochondrial membranes in aerobically grown wild-type cells. Anaerobically grown wild-type and all petite cultures contained cytochrome c peroxidase cytochemical reaction deposits on abundant cytoplasmic membranes and on the few mitochondrial profiles which also were seen in the electron photomicrographs. Biochemical studies corroborated the cytochemistry because mitochondrial fractions were greatly enriched in cytochrome c peroxidase activity for aerobically grown wild-type cultures, but petite and anaerobically grown wild-type cultures showed higher enzyme activities in supernatant fractions than was present in the corresponding particulate fractions after differential centrifugation. Evidence from low-temperature microspectroscopy, spectrophotometric assays of mitochondrial enzyme activities, and electron microscopy showed mitochondrial formation during the time required for preparation and lysis of spheroplasts from anaerobically grown cultures. The data were interpreted as indicating that cytochrome c peroxidase was an oxygen-inducible enzyme, and that there was a developmental relationship between enzyme-reactive membranes of mitochondria and cytoplasm during the period of respiratory adaptation.

Cell Membrane↗