structural modification
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- MeSH
- experimenty na zvířatech MeSH
- hemoglobiny účinky záření MeSH
- psi MeSH
- radiační účinky MeSH
- Check Tag
- psi MeSH
RNA polymerase II (RNA pol II) is not only the fundamental enzyme for gene expression but also the central coordinator of co-transcriptional processing. RNA pol II associates with a large number of enzymes and protein/RNA-binding factors through its C-terminal domain (CTD) that consists of tandem repeats of the heptapeptide consensus Y(1)S(2)P(3) T(4)S(5)P(6)S(7). The CTD is posttranslationally modified, yielding specific patterns (often called the CTD code) that are recognized by appropriate factors in coordination with the transcription cycle. Serine phosphorylations are currently the best characterized elements of the CTD code; however, the roles of the proline isomerization and other modifications of the CTD remain poorly understood. The dynamic remodeling of the CTD modifications by kinases, phosphatases, isomerases, and other enzymes introduce changes in the CTD structure and dynamics. These changes serve as structural switches that spatially and temporally regulate the binding of processing factors. Recent structural studies of the CTD bound to various proteins have revealed the basic rules that govern the recognition of these switches and shed light on the roles of these protein factors in the assemblies of the processing machineries.
- MeSH
- genetická transkripce MeSH
- methyltransferasy metabolismus MeSH
- peptidylprolylisomerasa metabolismus MeSH
- posttranslační úpravy proteinů * MeSH
- prolin metabolismus MeSH
- proteiny vázající RNA genetika metabolismus MeSH
- RNA-polymerasa II * chemie genetika metabolismus MeSH
- Saccharomyces cerevisiae enzymologie genetika MeSH
- sekvence aminokyselin MeSH
- terciární struktura proteinů MeSH
- transportní proteiny metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
BACKGROUND: Type I restriction-modification (R-M) systems are the most complex restriction enzymes discovered to date. Recent years have witnessed a renaissance of interest in R-M enzymes Type I. The massive ongoing sequencing programmes leading to discovery of, so far, more than 1 000 putative enzymes in a broad range of microorganisms including pathogenic bacteria, revealed that these enzymes are widely represented in nature. The aim of this study was characterisation of a putative R-M system EcoA0ORF42P identified in the commensal Escherichia coli A0 34/86 (O83: K24: H31) strain, which is efficiently used at Czech paediatric clinics for prophylaxis and treatment of nosocomial infections and diarrhoea of preterm and newborn infants. RESULTS: We have characterised a restriction-modification system EcoA0ORF42P of the commensal Escherichia coli strain A0 34/86 (O83: K24: H31). This system, designated as EcoAO83I, is a new functional member of the Type IB family, whose specificity differs from those of known Type IB enzymes, as was demonstrated by an immunological cross-reactivity and a complementation assay. Using the plasmid transformation method and the RM search computer program, we identified the DNA recognition sequence of the EcoAO83I as GGA(8N)ATGC. In consistence with the amino acids alignment data, the 3' TRD component of the recognition sequence is identical to the sequence recognized by the EcoEI enzyme. The A-T (modified adenine) distance is identical to that in the EcoAI and EcoEI recognition sites, which also indicates that this system is a Type IB member. Interestingly, the recognition sequence we determined here is identical to the previously reported prototype sequence for Eco377I and its isoschizomers. CONCLUSION: Putative restriction-modification system EcoA0ORF42P in the commensal Escherichia coli strain A0 34/86 (O83: K24: H31) was found to be a member of the Type IB family and was designated as EcoAO83I. Combination of the classical biochemical and bacterial genetics approaches with comparative genomics might contribute effectively to further classification of many other putative Type-I enzymes, especially in clinical samples.
- MeSH
- bakteriální proteiny genetika metabolismus MeSH
- DNA restrikčně-modifikační enzymy genetika metabolismus MeSH
- Escherichia coli enzymologie genetika MeSH
- financování organizované MeSH
- genomika MeSH
- proteiny z Escherichia coli genetika metabolismus MeSH
- protilátky bakteriální metabolismus MeSH
- restrikční endonukleasy typu I genetika metabolismus MeSH
- sekvence nukleotidů MeSH
- sekvenční homologie nukleových kyselin MeSH
- sekvenční seřazení MeSH
- testy genetické komplementace MeSH
Recently, we described the cold-dependent detection of an epitope, epiC, that was selectively recognized by a monoclonal anti-actin antibody at 4 degrees C, but not at RT, in the early replicating chromatin domains of human fibroblast cell nuclei and chromosomes. EpiC was present in a distinct cell cycle window extending from S-phase throughout mitosis until early G1-phase of the next cell generation, indicating its possible involvement in the transfer/maintenance of epigenetic information on transcriptionally competent parts of the genome. However, the molecular nature of epiC remained unresolved. Here we identified epiC as a dual post-translational modification on the same histone H4 tail, which was immunodetected for the first time. We show that the antibody selectively recognized a synthetic peptide of the histone H4 region K12-L22 containing acetylated K16 and dimethylated K20 (H4K16ac-K20me2) at 4 degrees C, but not at RT. Moreover, we show that the peptide containing acetylated K16 and either unmodified or monomethylated K20 was recognized by this antibody at both temperatures. The present and previous results together indicate that, by acetylation of histone H4 K16 during S-phase, the early replicating chromatin domains acquire the H4K16ac-K20me2 epigenetic label that persists on the chromatin throughout mitosis and become deacetylated during early G1-phase of the next cell cycle.
- MeSH
- acetylace MeSH
- buněčné linie MeSH
- buněčný cyklus fyziologie genetika MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- epigeneze genetická fyziologie genetika MeSH
- epitopy chemie imunologie MeSH
- histony imunologie metabolismus MeSH
- imunoblotting MeSH
- lidé MeSH
- metylace MeSH
- peptidy chemická syntéza chemie imunologie MeSH
- posttranslační úpravy proteinů MeSH
- teplota MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Our previous research on immunostimulatory properties of trilobolide and its structurally related natural analogues isolated from Laser trilobum (L.) Borkh., encouraged us to investigate structurally related guaianolides belonging to a specific group of sesquiterpene lactones with characteristic glycol moiety attached to the lactone ring. Ever increasing attention has been paid to certain guaianolides such as thapsigargin and trilobolide for their promising anti-inflammatory, anticancer, anti-infectious and SERCA inhibitory activities. However, due to their alkylation capabilities, they might be cytotoxic. Search for compounds with preserved immunobiological properties and decreased cytotoxicity led us to transform some of their structural features, particularly those related to their side chain functionality. For this reason, we prepared a series of over 20 various deacylated, acyl modified, or relactonized derivatives of trilobolide. The immunobiological effects were screened in vitro using the rat peritoneal cells primed with lipopolysaccharide. Secretion of interferon-γ (IFN-γ), interleukins (IL) IL-1β, IL-6 and tumour necrosis factor-α (TNF-α) were determined by ELISA, and nitric oxide (NO) production by Griess reagent. Relation between the molecular structure and immunobiological activity was investigated. Acetylation at 7-OH and 11-OH positions of the lactone ring, or acyl modification of the guaianolide functionalities (including relactonization) of trilobolide, led to inability to stimulate secretion of cytokines and production of NO. Interestingly, minor structural changes achieved by catalytic hydrogenation or hydrogenolysis retained the original immunoactivity of trilobolide. It can be concluded that several new chemically transformed sesquiterpene lactones resembling the immunobiological properties of trilobolide or thapsigargin were prepared and identified. The implication of the lactone vicinal diol (glycol) moiety, combined with other structure functionality, was confirmed as essential for immune properties of the trilobolide or thapsigargin type of guaianolides.
- MeSH
- Apiaceae chemie MeSH
- butyráty chemie MeSH
- cytokiny metabolismus MeSH
- furany chemie MeSH
- krysa rodu rattus MeSH
- kultivované buňky MeSH
- laktony chemie farmakologie MeSH
- molekulární struktura MeSH
- oxid dusnatý metabolismus MeSH
- peritoneální makrofágy účinky léků MeSH
- potkani Wistar MeSH
- seskviterpeny chemie farmakologie MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- Apiaceae chemie MeSH
- butyráty chemie farmakologie MeSH
- cytokiny imunologie MeSH
- furany chemie farmakologie MeSH
- kultivované buňky MeSH
- lidé MeSH
- molekulární struktura MeSH
- myši MeSH
- oxid dusnatý imunologie MeSH
- peritoneum cytologie MeSH
- potkani Wistar MeSH
- semena rostlinná chemie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Oxidative stress in humans is related to various pathophysiological processes, which can manifest in numerous diseases including cancer, cardiovascular diseases, and Alzheimer's disease. On the atomistic level, oxidative stress causes posttranslational modifications, thus inducing structural and functional changes into the proteins structure. This study focuses on fibrinogen, a blood plasma protein that is frequently targeted by reagents causing posttranslational modifications in proteins. Fibrinogen was in vitro modified by three reagents, namely sodium hypochlorite, malondialdehyde, and 3-morpholinosydnonimine that mimic the oxidative stress in diseases. Newly induced posttranslational modifications were detected via mass spectrometry. Electron microscopy was used to visualize changes in the fibrin networks, which highlight the extent of disturbances in fibrinogen behavior after exposure to reagents. We used molecular dynamics simulations to observe the impact of selected posttranslational modifications on the fibrinogen structure at the atomistic level. In total, 154 posttranslational modifications were identified, 84 of them were in fibrinogen treated with hypochlorite, 51 resulted from a reaction of fibrinogen with malondialdehyde, and 19 were caused by 3-morpholinosydnonimine. Our data reveal that the stronger reagents induce more posttranslational modifications in the fibrinogen structure than the weaker ones, and they extensively alter the architecture of the fibrin network. Molecular dynamics simulations revealed that the effect of posttranslational modifications on fibrinogen secondary structure varies from negligible alternations to serious disruptions. Among the serious disruptions is the oxidation of γR375 resulting in the release of Ca2+ ion that is necessary for appropriate fibrin fiber formation. Folding of amino acids γE72-γN77 into a short α-helix is a result of oxidation of γP76 to glutamic acid. The study describes behaviour of fibrinogen coiled-coil connecter in the vicinity of plasmin and hementin cleavage sites.
