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1.
Nanotechnology ; 33(49)2022 Sep 21.
Article in English | MEDLINE | ID: mdl-36049458

ABSTRACT

The self-ordered anodic aluminium oxide (AAO) structure consists of micron-scale domains-defect-free areas with a hexagonal arrangement of pores. A substantial increase in domain size is possible solely by pre-patterning the aluminium surface in the form of a defect-free hexagonal array of concaves, which guide the pore growth during subsequent anodization. Among the numerous pre-patterning techniques, direct etching by focused gallium ion beam (Ga FIB) allows the preparation of AAO with a custom-made geometry through precise control of the irradiation positions, beam energy, and ion dosage. The main drawback of the FIB approach includes gallium contamination of the aluminium surface. Here, we propose a multi-step anodizing procedure to prevent gallium incorporation into the aluminium substrate. The suggested approach successfully covers a wide range of AAO interpore distances from 100 to 500 nm. In particular, anodization of FIB pre-patterned aluminium in 0.1 M phosphoric acid at 195 V to prepare AAO with the interpore distance of about 500 nm was demonstrated for the first time. The quantification of the degree of pore ordering reveals the fraction of pores in hexagonal coordination above 96% and the in-plane mosaicity below 3° over an area of about 1000µm2. Large-scale defect-free AAO structures are promising for creating photonic crystals and hyperbolic metamaterials with distinct functional properties.

2.
Lett Appl Microbiol ; 67(3): 226-234, 2018 Sep.
Article in English | MEDLINE | ID: mdl-29927502

ABSTRACT

Streptomyces cinnamonensis A495 is a variant of the monensin producer which instead of the native polyether antibiotic gives rise to antibiotic and anti-tumour shunt-product premonensin. Through the supplementation of the fermentation medium with suitable precursors, premonensin can be derivatized via the incorporation of new-to-nature extender units into the biosynthetic machinery. Polyketide extender units require activation, typically in form of coenzyme A-thioesters. These are membrane impermeable and thus in the past an artificial mimic was employed. Here, we show the use and preliminary characterization of a highly substrate promiscuous new enzyme for the endogenous thioester formation in a Streptomyces strain. These intracellularly activated alternative extender units are significantly better incorporated into premonensin than the synthetically activated counterparts. SIGNIFICANCE AND IMPACT OF THE STUDY: Polyketide natural products are of enormous relevance in medicine. The hit-rate in finding active compounds for the potential treatment of various diseases among this substance family of microbial origin is high. However, most polyketides require derivatization to render them suitable for the application. Of relevance in this field is the incorporation of artificial substances into the biogenesis of polyketides, hampered by both the microbial metabolism and the complexity of the enzymes involved. This manuscript describes the straightforward and selective biosynthetic incorporation of synthetic substances into a reduced polyketide and showcases a promising new enzyme to aid this purpose.


Subject(s)
Anti-Bacterial Agents/biosynthesis , Bacterial Proteins/metabolism , Monensin/biosynthesis , Polyketide Synthases/metabolism , Streptomyces/metabolism , Bacterial Proteins/genetics , Biosynthetic Pathways , Enzyme Activation , Fermentation , Polyketide Synthases/genetics , Streptomyces/enzymology , Streptomyces/genetics
3.
Clin Microbiol Infect ; 24(8): 896-899, 2018 Aug.
Article in English | MEDLINE | ID: mdl-29217278

ABSTRACT

OBJECTIVES: Epidemiological studies have suggested an association between increased colistin use and selection for inherently colistin-resistant Enterobacteriaceae (ICRE). We aimed to evaluate whether colistin exposure is a risk factor for ICRE infection. METHODS: A matched 1:1 case-control study including patients recently hospitalized for ≥14 days with ICRE infection as cases matched with similar patients with a clinical isolate of a colistin-susceptible Enterobacteriaceae as controls was performed. Univariate analysis using McNemar test and multivariate analysis were conducted to explore risk factors for ICRE isolation, including colistin exposure 90 days before the positive culture. RESULTS: We included 446 patients, 223 cases and 223 controls matched for gender, age, department, year, source of culture and duration of hospitalization before positive culture. Colistin exposure was significantly associated with ICRE isolation in both univariate (14/223, 6.3% of cases versus 4/223, 1.8% of controls, p 0.031) and multivariate analyses (odds ratio 4.415, 95% CI 1.078-18.082). Curtailed functional capacity was a significant risk factor for ICRE as well. Exposure to other broad-spectrum antibiotics was associated with isolation of a colistin-susceptible pathogen. CONCLUSIONS: Exposure to colistin is associated with an increased risk of isolating an inherently colistin-resistant Enterobacteriaceae in patients with prolonged hospitalization. This should be taken into account while considering empirical therapy for such patients. Use of colistin should be judicious. The correlation between duration and magnitude of exposure and ICRE infection should be investigated in further studies.


Subject(s)
Anti-Bacterial Agents/therapeutic use , Colistin/therapeutic use , Drug Resistance, Bacterial , Enterobacteriaceae Infections/drug therapy , Enterobacteriaceae Infections/microbiology , Enterobacteriaceae/drug effects , Aged , Aged, 80 and over , Anti-Bacterial Agents/pharmacology , Case-Control Studies , Colistin/pharmacology , Comorbidity , Enterobacteriaceae/genetics , Enterobacteriaceae Infections/epidemiology , Female , Humans , Male , Middle Aged , Retrospective Studies , Risk Factors , Treatment Outcome
4.
Org Biomol Chem ; 14(32): 7671-5, 2016 Aug 10.
Article in English | MEDLINE | ID: mdl-27452503

ABSTRACT

The controlled derivatization of natural products is of great importance for their use in drug discovery. The ideally rapid generation of compound libraries for structure-activity relationship studies is of particular concern. We here use modified biosynthesis for the generation of such a library of reduced polyketides to interfere with the oncogenic KRas pathway. The polyketide is derivatized via side chain alteration, and variations in its redox pattern and in its backbone chain length through manipulation in the corresponding polyketide synthase. Structural and biophysical analyses revealed the nature of the interaction between the polyketides and KRas-interacting protein PDE6δ. Non-natural polyketides with low nanomolar affinity to PDE6δ were identified.


Subject(s)
Polyketide Synthases/metabolism , Polyketides/metabolism , Humans , Polyketide Synthases/chemistry , Polyketides/chemistry , Proto-Oncogene Proteins p21(ras)/chemistry , Proto-Oncogene Proteins p21(ras)/metabolism , Structure-Activity Relationship
5.
Vasc Health Risk Manag ; 9: 535-49, 2013.
Article in English | MEDLINE | ID: mdl-24109189

ABSTRACT

BACKGROUND: The aim of this study was to compare the antihypertensive efficacy of losartan 100 mg + hydrochlorothiazide (HCTZ) 25 mg versus bisoprolol 10 mg + HCTZ 25 mg and their influence on arterial stiffness and central blood pressure (BP). METHODS: Of 60 patients with a mean BP of 173.3 ± 1.7/98.4 ± 1.2 mmHg, 59 were randomized to losartan + HCTZ (n = 32) or bisoprolol + HCTZ (n = 27). Amlodipine was added if target BP was not achieved at 1 month, and doxazosin was added if target BP was not achieved after 3 months. Body mass index, office and 24-hour ambulatory BP, pulse wave velocity (carotid-femoral [PWVE] and radial [PWVM]), noninvasive central systolic BP, augmentation index (AIx), laboratory investigations, and electrocardiography were done at baseline and after 6 months of treatment. RESULTS: Losartan + HCTZ was as effective as bisoprolol + HCTZ, with target office BP achieved in 96.9% and 92.6% of patients and target 24-hour BP in 75% and 66.7% of patients, respectively, after 6 months. Effective treatment of BP led to significant lowering of central systolic BP, but this was decreased to a significantly (P < 0.05) greater extent by losartan + HCTZ (-23.0 ± 2.3 mmHg) than by bisoprolol + HCTZ (-15.4 ± 2.9 mmHg) despite equal lowering of brachial BP. Factors correlated with central systolic BP and its lowering differed between the treatment groups. Losartan + HCTZ did not alter arterial stiffness patterns significantly, but bisoprolol + HCTZ significantly increased AIx. We noted differences in ΔPWVE, ΔPWVM, and ΔAIx between the groups in favor of losartan + HCTZ. Decreased heart rate was associated with higher central systolic BP and AIx in the bisoprolol + HCTZ group, but was not associated with increased AIx in the losartan + HCTZ group. CONCLUSION: Although both treatments decreased both office and 24-hour BP, losartan + HCTZ significantly decreased central systolic BP and had a more positive influence on pulse wave velocity, with a less negative effect of decreased heart rate on AIx and central systolic BP.


Subject(s)
Adrenergic beta-1 Receptor Antagonists/administration & dosage , Angiotensin II Type 1 Receptor Blockers/administration & dosage , Antihypertensive Agents/administration & dosage , Bisoprolol/administration & dosage , Blood Pressure/drug effects , Diuretics/administration & dosage , Hydrochlorothiazide/administration & dosage , Hypertension/drug therapy , Losartan/administration & dosage , Amlodipine/administration & dosage , Blood Pressure Monitoring, Ambulatory , Calcium Channel Blockers/administration & dosage , Chi-Square Distribution , Doxazosin/administration & dosage , Drug Combinations , Drug Therapy, Combination , Electrocardiography , Female , Heart Rate/drug effects , Humans , Hypertension/diagnosis , Hypertension/physiopathology , Male , Middle Aged , Pulse Wave Analysis , Severity of Illness Index , Time Factors , Treatment Outcome , Ukraine , Vascular Stiffness/drug effects
6.
Stomatologiia (Mosk) ; 92(2): 57-9, 2013.
Article in Russian | MEDLINE | ID: mdl-23715457

ABSTRACT

Effect of fluorides in drinking water on fluorine content in breast milk, the food for infants of the 1-sty year of life, was investigated. On determining fluorine concentration in urine and its excretion, fluorine intake by the infants was calculated under various alternatives (breast, mixed and artificial) of feeding. It has been found the in mixed and especially in artificial feeding, fluorine intake by the infants acquires uncontrollable character exceeding several times the dose of fluorine intake with breast milk under natural feeding. That was predominantly fluorine of drinking water. Mathematical formula for calculation of fluorine content in the food cooked for feeding of infants was elaborated. A computer program was formed be means of which calculation, control and management of fluorine intake are carried out in feeding alteration.


Subject(s)
Fluorides/standards , Fluorosis, Dental/epidemiology , Fluorosis, Dental/prevention & control , Breast Feeding , Child, Preschool , Fluorides/administration & dosage , Fluorides/adverse effects , Fluorosis, Dental/etiology , Humans , Infant , Milk, Human/chemistry , Nutritional Support/adverse effects , Nutritional Support/standards , Risk , Software
7.
Stomatologiia (Mosk) ; 90(1): 65-7, 2011.
Article in Russian | MEDLINE | ID: mdl-21378727

ABSTRACT

Fluoride content in drinking water, breast milk, cow milk, additional food for newborns prepared with water containing different amount of fluoride was determined. Fluoride excretion in urine since the first days of birth and up to 4 months of postnatal development was investigated in breast and artificially fed infants. When a neonate was fed with breast milk, fluoride was received in the amount no more than 20 mkg/day. The additional food contained fluoride which water mainly had. Water with high level of fluorine increased its content in the additional food up to the values not comparable to those in breast milk that presented danger of dental fluorosis development. Data on fluorine content in drinking water were absolutely necessary to calculate daily fluorides consumption by infants and to prevent dental fluorosis.


Subject(s)
Fluorides/administration & dosage , Fluorides/analysis , Fluorosis, Dental/prevention & control , Animals , Female , Fluorides/urine , Humans , Infant , Milk/chemistry , Milk Substitutes/chemistry , Milk, Human/chemistry , Russia , Water/chemistry , Water Supply/analysis
8.
Microbiology (Reading) ; 151(Pt 1): 281-290, 2005 Jan.
Article in English | MEDLINE | ID: mdl-15632445

ABSTRACT

The gene lndI is involved in the pathway-specific positive regulation of biosynthesis of the antitumour polyketide landomycin E in Streptomyces globisporus 1912. LndI was overexpressed in Escherichia coli as a protein C-terminally fused to the intein-chitin-binding-domain tag and purified in a one-step column procedure. Results of in vivo LndI titration, DNA gel mobility-shift assays and promoter-probing experiments indicate that LndI is an autoregulatory DNA-binding protein that binds to its own gene promoter and to the promoter of the structural gene lndE. Enhanced green fluorescent protein was used as a reporter to study the temporal and spatial pattern of lndI transcription. Expression of lndI started before cells entered mid-exponential phase and peak expression coincided with maximal accumulation of landomycin E and biomass. In solid-phase analysis, lndI expression was evident in substrate mycelia but was absent from aerial hyphae and spores.


Subject(s)
Aminoglycosides/biosynthesis , Bacterial Proteins/metabolism , DNA-Binding Proteins/metabolism , Streptomyces/metabolism , Up-Regulation , Bacterial Proteins/genetics , DNA-Binding Proteins/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression Regulation, Bacterial , Microscopy, Confocal , Molecular Sequence Data , Promoter Regions, Genetic , Recombinant Fusion Proteins/metabolism , Sequence Analysis, DNA , Streptomyces/genetics , Streptomyces/ultrastructure , Transcription, Genetic
9.
Prikl Biokhim Mikrobiol ; 39(6): 652-5, 2003.
Article in Russian | MEDLINE | ID: mdl-14714478

ABSTRACT

Effects of co-cultivation of higher Basidiomycetes and Phycomycetes on the biosynthesis of cellulases, amylases, and proteases was studied. Four optimal pairs of fungal cultures were selected. Of these, three pairs belonged to higher fungi, and one pair was constituted by fungi of distinct ecological groups, a macromycete and a micromycete. The activities of amylase and protease were 1.5 to 2 times higher, and the activity of cellulase was lower during the growth of higher fungi associations. The mixed association of the macromycete Schizophyllum commune and the micromycete Mucor sp. was the most active producer of hydrolytic enzymes. During the growth of this mixed association, a fourfold and 1.5-fold increases were observed in the activity of endoglucanase and protease, respectively, paralleled by stimulation of amylase formation.


Subject(s)
Amylases/biosynthesis , Basidiomycota/enzymology , Cellulases/biosynthesis , Endopeptidases/biosynthesis , Mitosporic Fungi/enzymology , Mucor/enzymology , Basidiomycota/growth & development , Coculture Techniques , Hydrolysis , Mitosporic Fungi/growth & development , Mucor/growth & development
10.
Plant J ; 28(3): 245-55, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11722768

ABSTRACT

Poly(ADP-ribos)ylation is a reversible protein modification that in higher plants is catalyzed by two structurally different poly(ADP-ribose) polymerases, App and Zap. In vivo imaging of green-fluorescent protein (GPF) fusions showed that both Zap and App were associated with chromatin through the cell cycle progression. The in vivo behaviour of the App-GFP protein fusions can be attributed to the activity of two NASA motifs that mediate protein-protein interactions and nucleic acid binding. Expression of Zap deletion variants revealed that both Zn fingers and helix-turn-helix domains contributed to the association with chromosomes, whereas the localization in the nucleoplasm was mostly determined by the Zn fingers. The results highlight novel properties of protein sequences found in plant poly(ADP-ribose) polymerases and suggest important functions for this class of nuclear enzymes in chromosome dynamics.


Subject(s)
Arabidopsis/enzymology , Plant Proteins/genetics , Poly(ADP-ribose) Polymerases/metabolism , Amino Acid Sequence , Arabidopsis/genetics , Cell Culture Techniques , Chromatin , Cloning, Molecular , Genetic Vectors , Green Fluorescent Proteins , Luminescent Proteins , Microscopy, Confocal , Molecular Sequence Data , Plant Proteins/chemistry , Poly Adenosine Diphosphate Ribose/chemistry , Poly(ADP-ribose) Polymerases/chemistry , Protein Structure, Tertiary , Recombinant Fusion Proteins , Sequence Analysis, DNA , Sequence Analysis, Protein
12.
Plant Cell ; 13(1): 89-100, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11158531

ABSTRACT

A mutation in the Arabidopsis gene STARIK leads to dwarfism and chlorosis of plants with an altered morphology of leaf and cell nuclei. We show that the STARIK gene encodes the mitochondrial ABC transporter Sta1 that belongs to a subfamily of Arabidopsis half-ABC transporters. The severity of the starik phenotype is suppressed by the ectopic expression of the STA2 homolog; thus, Sta1 function is partially redundant. Sta1 supports the maturation of cytosolic Fe/S protein in Deltaatm1 yeast, substituting for the ABC transporter Atm1p. Similar to Atm1p-deficient yeast, mitochondria of the starik mutant accumulated more nonheme, nonprotein iron than did wild-type organelles. We further show that plant mitochondria contain a putative l-cysteine desulfurase. Taken together, our results suggest that plant mitochondria possess an evolutionarily conserved Fe/S cluster biosynthesis pathway, which is linked to the intracellular iron homeostasis by the function of Atm1p-like ABC transporters.


Subject(s)
Arabidopsis Proteins , Arabidopsis/genetics , Iron/metabolism , Mutation , Plant Proteins/genetics , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis/physiology , Cell Nucleus/ultrastructure , Gene Expression Profiling , Iron-Sulfur Proteins/biosynthesis , Mitochondria/metabolism , Plant Leaves/anatomy & histology
13.
J Exp Bot ; 51(351): 1761-2, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11053466

ABSTRACT

Budding yeast strains that produced the Arabidopsis thaliana protein CEF or its amino-terminal proline-rich domain were more tolerant to hydroperoxides. CEF is homologous to animal and yeast Sec24 proteins. These data suggest that CEF plays a protective role through protein transport during growth under pro-oxidant conditions.


Subject(s)
Arabidopsis/genetics , Membrane Proteins/physiology , Oxidative Stress , Plant Proteins/physiology , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae/physiology , Amino Acid Sequence , Membrane Proteins/genetics , Molecular Sequence Data , Plant Proteins/genetics , Saccharomyces cerevisiae/genetics , Sequence Homology, Amino Acid , Vesicular Transport Proteins
14.
FEBS Lett ; 482(1-2): 19-24, 2000 Sep 29.
Article in English | MEDLINE | ID: mdl-11018516

ABSTRACT

The Saccharomyces cerevisiae strain WYT, deficient in the YAP1 transcription factor, was used in a molecular screen to identify genes from Arabidopsis thaliana that could overcome the oxidative stress-sensitive phenotype of these yeast cells. A cDNA named CEO1 increased the tolerance to oxidative damage caused by tert-butylhydroperoxide of both the Yap1(-) mutant and the wild-type yeast. Additionally, in Yap1(-) yeast, CEO1 also induced cross-tolerance to oxidative damage caused by hydrogen peroxide and diamide. CEO1 was assigned as being part of a small gene family that, until now, is exclusively restricted to plants. In Arabidopsis, CEO1 was produced in all organs, especially in roots and stems. By using the yeast two-hybrid system, proteins that specifically interact with CEO1 in yeast were identified, and putative DNA-binding proteins were consistently recovered.


Subject(s)
Arabidopsis Proteins , Arabidopsis/genetics , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae/genetics , Transcription Factors/genetics , Transcription Factors/metabolism , Amino Acid Sequence , Cloning, Molecular , DNA-Binding Proteins/genetics , Escherichia coli , Fungal Proteins/genetics , Molecular Sequence Data , Multigene Family , Oxidative Stress , Plant Proteins/chemistry , Plant Proteins/genetics , Plant Proteins/metabolism , Plant Roots/metabolism , Plant Stems/metabolism , Protein Isoforms/chemistry , Protein Isoforms/genetics , Recombinant Proteins/metabolism , Saccharomyces cerevisiae/physiology , Sequence Alignment , Sequence Homology, Amino Acid , Transcription Factors/chemistry
15.
Eur J Biochem ; 267(12): 3661-71, 2000 Jun.
Article in English | MEDLINE | ID: mdl-10848984

ABSTRACT

The zeta-crystallin (ZCr) gene P1 of Arabidopsis thaliana, known to confer tolerance toward the oxidizing drug 1,1'-azobis(N, N-dimethylformamide) (diamide) to yeast [Babiychuk, E., Kushnir, S., Belles-Boix, E., Van Montagu, M. & Inzé, D. (1995) J. Biol. Chem. 270, 26224], was expressed in Escherichia coli to characterize biochemical properties of the P1-zeta-crystallin (P1-ZCr). Recombinant P1-ZCr, a noncovalent dimer, showed NADPH:quinone oxidoreductase activity with specificity to quinones similar to that of guinea-pig ZCr. P1-ZCr also catalyzed the divalent reduction of diamide to 1,2-bis(N,N-dimethylcarbamoyl)hydrazine, with a kcat comparable with that for quinones. Two other azodicarbonyl compounds also served as substrates of P1-ZCr. Guinea-pig ZCr, however, did not catalyze the azodicarbonyl reduction. Hence, plant ZCr is distinct from mammalian ZCr, and can be referred to as NADPH:azodicarbonyl/quinone reductase. The quinone-reducing reaction was accompanied by radical chain reactions to produce superoxide radicals, while the azodicarbonyl-reducing reaction was not. Specificity to NADPH, as judged by kcat/Km, was > 1000-fold higher than that to NADH both for quinones and diamide. N-Ethylmaleimide and p-chloromercuribenzoic acid inhibited both quinone-reducing and diamide-reducing activities. Both NADPH and NADP+ suppressed the inhibition, but NADH did not, suggesting that sulfhydryl groups reside in the binding site for the phosphate group on the adenosine moiety of NADPH. The diamide-reducing activity of P1-ZCr accounts for the tolerance of P1-overexpressing yeast to diamide. Other possible physiological functions of P1-ZCr in plants are discussed.


Subject(s)
Arabidopsis/metabolism , Crystallins/metabolism , Diamide/metabolism , Oxidoreductases/metabolism , Plant Proteins/metabolism , Amino Acid Sequence , Animals , Crystallins/drug effects , Crystallins/genetics , Dicumarol/pharmacology , Dimerization , Enzyme Inhibitors/pharmacology , Escherichia coli/genetics , Guinea Pigs , Hydrogen-Ion Concentration , Molecular Sequence Data , NADP/metabolism , Nitrofurantoin/pharmacology , Nucleotides/metabolism , Oxidative Stress , Plant Proteins/drug effects , Plant Proteins/genetics , Quinone Reductases/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Substrate Specificity , Sulfhydryl Compounds/chemistry
16.
Plant Physiol ; 118(3): 1005-14, 1998 Nov.
Article in English | MEDLINE | ID: mdl-9808745

ABSTRACT

Ascorbate peroxidases are important enzymes that detoxify hydrogen peroxide within the cytosol and chloroplasts of plant cells. To better understand their role in oxidative stress tolerance, the transcriptional regulation of the apx1 gene from Arabidopsis was studied. The apx1 gene was expressed in all tested organs of Arabidopsis; mRNA levels were low in roots, leaves, and stems and high in flowers. Steady-state mRNA levels in leaves or cell suspensions increased after treatment with methyl viologen, ethephon, high temperature, and illumination of etiolated seedlings. A putative heat-shock cis element found in the apx1 promoter was shown to be recognized by the tomato (Lycopersicon esculentum) heat-shock factor in vitro and to be responsible for the in vivo heat-shock induction of the gene. The heat-shock cis element also contributed partially to the induction of the gene by oxidative stress. By using in vivo dimethyl sulfate footprinting, we showed that proteins interacted with a G/C-rich element found in the apx1 promoter.


Subject(s)
Arabidopsis/genetics , Heat-Shock Proteins/metabolism , Peroxidases/genetics , Promoter Regions, Genetic , Arabidopsis/enzymology , Ascorbate Peroxidases , Base Sequence , DNA Footprinting , DNA Primers , DNA, Complementary , Oxidative Stress , RNA, Messenger/genetics , RNA, Messenger/metabolism , Sequence Homology, Nucleic Acid
17.
Plant J ; 15(5): 635-45, 1998 Sep.
Article in English | MEDLINE | ID: mdl-9778846

ABSTRACT

One of the immediate reactions of the mammalian cell to many environmental stresses is a massive synthesis of poly(ADP-ribose), catalyzed by poly(ADP-ribose) polymerase (PARP). Most of the biological functions attributed to PARP are inferred from experimentation with mammalian cells. In plants, the biology of PARP may be more complicated and diverse than was previously thought. Two poly(ADP-ribose) polymerase homologues were found in plants, the classical Zn-finger-containing polymerase (ZAP) and the structurally non-classical PARP proteins (APP and NAP), which lack the characteristic N-terminal Zn-finger domain. By enzymatic and cytological experiments the recombinant APP protein was shown to be located in the nucleus and to possess DNA-dependent poly(ADP-ribose) polymerase activity in yeast. The nuclear localization was further confirmed by the analysis of transgenic tobacco plants that expressed a translational gene fusion between APP and the bacterial beta-glucuronidase. The app promoter was transcriptionally up-regulated in cells pre-determined to die because of deficiency in a DNA ligase I.


Subject(s)
Arabidopsis/enzymology , Plant Proteins/genetics , Poly(ADP-ribose) Polymerases/genetics , Zea mays/enzymology , Amino Acid Sequence , Arabidopsis/genetics , DNA Ligases/metabolism , Escherichia coli , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Plant , Molecular Sequence Data , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Plant Proteins/chemistry , Poly(ADP-ribose) Polymerases/chemistry , Saccharomyces cerevisiae , Zea mays/genetics , Zinc Fingers
18.
Plant Mol Biol ; 38(3): 491-6, 1998 Oct.
Article in English | MEDLINE | ID: mdl-9747856

ABSTRACT

Transgenic maize (Zea mays L.) and tobacco (Nicotiana tabacum Petit Havana SR1) plants have been generated, which overproduce a mitochondrial Nicotiana plumbaginifolia manganese superoxide dismutase (MnSOD) in chloroplasts. For this, the mature MnSOD-coding sequence was fused to a chloroplast transit peptide from a Pisum sativum ribulose-1,5-bisphosphate carboxylase (Rubisco) gene and expression of the chimeric gene was driven by the cauliflower mosaic virus (CaMV) 35S promoter. The transgenic MnSOD gene product was correctly targeted to the chloroplasts both in maize and tobacco. However, despite the use of the CaMV 35S promoter, the MnSOD was predominantly localized in the chloroplasts of the bundle sheath cells of maize. Furthermore, the transit peptide was cleaved off at a different position in maize and tobacco.


Subject(s)
Nicotiana/metabolism , Plant Proteins/metabolism , Plants, Toxic , Zea mays/metabolism , Amino Acid Sequence , Base Sequence , Binding Sites/genetics , Caulimovirus/genetics , Chloroplasts/metabolism , DNA, Recombinant/genetics , Gene Expression , Immunohistochemistry , Molecular Sequence Data , Plant Proteins/genetics , Plants, Genetically Modified , Promoter Regions, Genetic , Protein Processing, Post-Translational , Protein Sorting Signals/genetics , Protein Sorting Signals/metabolism , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Ribulose-Bisphosphate Carboxylase/genetics , Ribulose-Bisphosphate Carboxylase/metabolism , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism , Nicotiana/genetics , Nicotiana/virology , Zea mays/genetics , Zea mays/virology
19.
Article in Russian | MEDLINE | ID: mdl-9575622

ABSTRACT

Peculiarities of paroxysmal course of autonomic dystonia of cardiac type were studied in 148 juveniles aged 14-15 years. It was found that frequency of panic attacks was very high, i.e. they occurred in 67-92.4% of the cases in dependence on severity of the disease. Panic attacks had no definite clinical picture at the onset. However, as the intensity of the main clinical manifestation of the disease (chest-pain syndrome) increased they acquired clear-cut sympatho-adrenal or vago-insular direction. Short duration and incomplete nels are typical for panic attacks in these patients.


Subject(s)
Autonomic Nervous System Diseases/complications , Chest Pain/complications , Dystonia/complications , Panic Disorder/diagnosis , Adolescent , Female , Humans , Male , Panic Disorder/etiology , Syndrome , Time Factors
20.
Proc Natl Acad Sci U S A ; 94(23): 12722-7, 1997 Nov 11.
Article in English | MEDLINE | ID: mdl-9356517

ABSTRACT

Large quantities of DNA sequence information about plant genes are rapidly accumulating in public databases, but to progress from DNA sequence to biological function a mutant allele for each of the genes ideally should be available. Here we describe a gene trap construct that allowed us to disrupt transcribed genes with a high efficiency in Arabidopsis thaliana. In the T-DNA vector used, the expression of a bacterial reporter gene coding for neomycin phosphotransferase II (nptII) depends on the in vivo generation of a translation fusion upon the T-DNA integration into the Arabidopsis genome. Analysis of 20 selected transgenic lines showed that 12 lines are T-DNA insertion mutants. The disrupted genes analyzed encoded ribosomal proteins (three lines), aspartate tRNA synthase, DNA ligase, basic-domain leucine zipper DNA binding protein, ATP-binding cassette transporter, and five proteins of unknown function. Four tagged genes were new for Arabidopsis. The results presented here suggest that gene trapping, using nptII as a reporter gene, can be as high as 80% and opens novel perspectives for systematic gene tagging in A. thaliana.


Subject(s)
Arabidopsis/genetics , DNA, Plant , Gene Transfer Techniques , Genes, Plant , Mutagenesis, Insertional , Genetic Vectors , Molecular Sequence Data
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