Functional monomers
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The polarity of (poly)methacrylate monolithic capillary columns was varied by using alkylmethacrylate monomers with butyl, cyclohexyl, 2-ethylhexyl, lauryl, and stearyl functional groups in the polymerization mixture. The hydrodynamic properties, as well as the retention characteristics in RP-LC of small molecules (alkylbenzenes) and of proteins under gradient elution conditions were studied. The RP selectivity depends on the type of alkyl chain in methacrylate monomer; however, there was no direct correlation between the size of the monomer molecule and methylene or aromatic selectivity of the monlithic column. The lowest selectivity was found for column based on lauryl methacrylate monomer. On the other hand, butyl methacrylate column shows high phenyl selectivity and the column with stearyl methacrylate possesses the highest methylene selectivity for small molecules. The retention increases with longer alkyl chain in methacrylate monomer, especially for high molar mass proteins on all prepared columns and showed gradient elution behavior of proteins in agreement with the linear solvent strength gradient model. The poly(laurylmethacrylate) column showed lowest hydrophobicity but best efficiency for proteins of all columns tested.
In addition to their force-generating motor domains, kinesin motor proteins feature various accessory domains enabling them to fulfill a variety of functions in the cell. Human kinesin-3, Kif14, localizes to the midbody of the mitotic spindle and is involved in the progression of cytokinesis. The specific motor properties enabling Kif14's cellular functions, however, remain unknown. Here, we show in vitro that the intrinsically disordered N-terminal domain of Kif14 enables unique functional diversity of the kinesin. Using single molecule TIRF microscopy, we found that Kif14 exists either as a diffusible monomer or as processive dimer and that the disordered domain (1) enables diffusibility of the monomeric Kif14, (2) renders the dimeric Kif14 super-processive and enables the kinesin to pass through highly crowded areas, (3) enables robust, autonomous Kif14 tracking of growing microtubule tips, independent of microtubule end-binding (EB) proteins, and (4) is sufficient to enable crosslinking of parallel microtubules and necessary to enable Kif14-driven sliding of antiparallel ones. We explain these features of Kif14 by the observed diffusible interaction of the disordered domain with the microtubule lattice and the observed increased affinity of the disordered domain for GTP-bound tubulin. We suggest that the disordered domain tethers the motor domain to the microtubule providing a diffusible foothold and a regulatory hub, tuning the kinesin's interaction with microtubules. Our findings thus exemplify pliable protein tethering as a fundamental mechanism of molecular motor regulation.
- MeSH
- aparát dělícího vřeténka fyziologie MeSH
- cytokineze * MeSH
- kineziny chemie genetika metabolismus MeSH
- lidé MeSH
- mikrotubuly metabolismus MeSH
- onkogenní proteiny chemie genetika metabolismus MeSH
- vazba proteinů MeSH
- vnitřně neuspořádané proteiny chemie genetika metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The chain coordination polymerization of (ethynylarene)carbaldehydes with unprotected carbaldehyde groups, namely ethynylbenzaldehydes, 1-ethynylbenzene-3,5-dicarboxaldehyde, and 3-[(4-ethynylphenyl)ethynyl]benzaldehyde, is reported for the first time. Polymerization is catalyzed with various Rh(I) catalysts and yields poly(arylacetylene)s with one or two pendant carbaldehyde groups per monomeric unit. Surprisingly, the carbaldehyde groups of the monomers do not inhibit the polymerization unlike the carbaldehyde group of unsubstituted benzaldehyde that acts as a strong inhibitor of Rh(I) catalyzed polymerization of arylacetylenes. The inhibition ability of carbaldehyde groups in (ethynylarene)carbaldehydes seems to be eliminated owing to a simultaneous presence of unsaturated ethynyl groups in (ethynylarene)carbaldehydes. The reactive carbaldehyde groups make poly[(ethynylarene)carbaldehyde]s promising for functional appreciation via various postpolymerization modifications. The introduction of photoluminescence or chirality to poly(ethynylbenzaldehyde)s via quantitative modification of their carbaldehyde groups in reaction with either photoluminescent or chiral primary amines under formation of the polymers with Schiff-base-type pendant groups is given as an example.
- MeSH
- aldehydy chemie MeSH
- benzaldehydy chemie MeSH
- chemické modely MeSH
- fluorescenční spektrometrie MeSH
- katalýza MeSH
- molekulární struktura MeSH
- polyacetyleny chemická syntéza chemie MeSH
- polymerizace * MeSH
- polymery chemická syntéza chemie MeSH
- protonová magnetická rezonanční spektroskopie MeSH
- Publikační typ
- časopisecké články MeSH
14-3-3 proteins are universal regulatory proteins and their function depends on their oligomeric form which may alter between the monomeric, homodimeric and heterodimeric states. The populations of individual oligomeric forms are controlled by Kd values of the dimer-monomer equilibria between the involved isoforms. This complex picture is extended by post-translational modifications, e.g. phosphorylation. In this work, we describe the equilibria between monomers, homo- and heterodimers of the 14-3-3ζ isoform in the unmodified and phosphorylated form. To cover a wide range of dimerization affinities, we combined solution NMR, microscale thermophoresis, native PAGE, and a set of novel fluorescence assays. Using a FRET based assay, we also determined the kinetic parameters of dimerization. We found that phosphorylation of 14-3-3ζ at Ser58 increases its homodimeric Kd value by 6 orders of magnitude. The presented assays allow to efficiently monitor 14-3-3ζ dimerization as a function of external factors, such as temperature, salt concentration, and client protein binding. For instance, we obtained values of both transient and equilibrium thermodynamic constants for the dimerization, and observed a substantial decrease of 14-3-3ζ dimer dissociation rate upon binding to the doubly phosphorylated regulatory domain of tyrosine hydroxylase. In summary, our work provides a conceptual framework to characterise the isoform exchanges of homo- and heterodimers which can significantly deepen our knowledge about the regulatory function of 14-3-3 proteins.
- MeSH
- fosforylace MeSH
- lidé MeSH
- multimerizace proteinu MeSH
- proteiny 14-3-3 * chemie MeSH
- termodynamika MeSH
- vazba proteinů MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Polymers with lower critical solution temperature (LCST) are molecularly soluble in their solutions at low temperatures but, when heated above their cloud point temperature, these polymers assemble into supramolecular particles or macroscopic precipitates. These particles or precipitated polymeric depots can be used for diagnostics, targeted drug delivery, controlled drug release from a depot or a combination of diagnostics and therapy (theranostics). Herein, we describe smart polymer systems which contain N-(2,2-difluoroethyl)acrylamide monomer unit (DFEA) and form polymeric nanoparticles upon heating or precipitate after injection to polymeric implants/depots. Due to a high fluorine content and to relaxation properties of this element, these polymers are suitable as tracers for the 19F MRI method, a promising non-invasive diagnostic tool. Moreover, DFEA copolymers can contain monomers that react to additional physicochemical properties, resulting in multiresponsive polymers (pH- or redox-responsive), which can be used for smart drug delivery systems with controlled release of drugs in the target environment.
- MeSH
- dichlorethyleny MeSH
- kontrastní látky * MeSH
- lidé MeSH
- magnetická rezonanční tomografie MeSH
- Check Tag
- lidé MeSH
G-quadruplexes are noncanonical nucleic acid structures formed from stacked guanine tetrads. They are frequently used as building blocks and functional elements in fields such as synthetic biology and also thought to play widespread biological roles. G-quadruplexes are often studied as monomers, but can also form a variety of higher-order structures. This increases the structural and functional diversity of G-quadruplexes, and recent evidence suggests that it could also be biologically important. In this review, we describe the types of multimeric topologies adopted by G-quadruplexes and highlight what is known about their sequence requirements. We also summarize the limited information available about potential biological roles of multimeric G-quadruplexes and suggest new approaches that could facilitate future studies of these structures.
Magnetic resonance imaging (MRI) using 19F-based tracers has emerged as a promising multi-purpose noninvasive diagnostic tool and its application requires the use of various 19F-based tracers for the intended diagnostic purpose. In this study, we report a series of double-stimuli-responsive polymers for use as injectable implants, which were designed to form implants under physiological conditions, and to subsequently dissolve with different dissolution rates (t1/2 ranges from 30 to more than 250 days). Our polymers contain a high concentration of fluorine atoms, providing remarkable signal detectability, and both a hydrophilic monomer and a pH-responsive monomer that alter the biodistribution properties of the implant. The implant location and dissolution were observed using 19F MRI, which allows the anatomic extent of the implant to be monitored. The dissolution kinetics and biocompatibility of these materials were thoroughly analyzed. No sign of toxicity in vitro or in vivo or pathology in vivo was observed, even in chronic administration. The clinical applicability of our polymers was further confirmed via imaging of a rat model by employing an instrument currently used in human medicine.
Východisko: Mnohočetná angiomatóza je raritní onemocnění s postižením více orgánů a tkání angiomatózními útvary s rizikem smrtelného krvácení. Popis případu: V popisovaném případu byly postiženy kosti, pleurální i peritoneální dutina a trávicí trubice. Pacient dlouhodobě dostává zoledronát, již po druhé dávce zoledronátu vymizely bolesti kostní. Účinnost antiangiogenních léků jsme hodnotili dle CT a MR zobrazení. Protože angiomatózní proliferace je spojena s chronickou diseminovanou intravaskulární koagulací (DIC) a anemizací, hodnotíme léčebnou odpověď také dle krevního obrazu a dle koncentrace fibrinogenu, D-dimerů a průkazu solubilních fibrinových monomerů. Výsledky: Před léčbou byly hodnoty D-dimerů vyšší než 20 μg/ml, fibrinogen 1,4 g/l a byly přítomny solubilní fibrinové monomery. Při léčbě interferonem α 6 milionů j. 3krát týdně s redukcí dávky po 10 měsících se zvýšil medián koncentrace fibrinogenu na 1,5 (1,2–2,0) g/l, poklesl medián hodnoty D-dimerů na 17,2 (13,4–20,0) μg/ml, stále byly přítomny fibrinové monomery. Při terapii thalidomidem (100 mg/den) poklesl medián D-dimerů na 6,07 (4,71–10,21) μg/ml a zvýšil se medián hodnoty fibrinogenu na 1,9 g/l, solubilní fibrinové monomery nebyly průkazné. Dle CT zobrazení se výrazně zmenšily angiomatózní masy. Progredující neuropatie byla důvodem pro snížení dávky thalidomidu na 50 mg/den, při níž se ale zvýšily D-dimery. Při podávání lenalidomidu 10 mg/den se zvýšil medián koncentrace D-dimerů na 10,8 (10,8–17,35) a poklesla hodnota hemoglobinu na medián 124 (135–117) g/l. Opět byl pozitivní průkaz solubilních fibrinových monomerů. Proto jsme zkombinovali nízkou dávku lenalidomidu 10 mg/den s thalidomidem 100 mg a posléze 50 mg denně. Při léčbě lenalidomidem 10 mg a thalidomidem 50 mg byl medián hodnoty D-dimerů 9,32 a onemocnění bylo po 9 měsíců stabilní. Závěr: Thalidomid 100 mg/den stabilizoval mnohočetnou angiomatózu lépe než interferon alfa. Thalidomid 50 mg/den již neudržel stabilitu nemoci. Lenalidomid v dávce 10 mg byl excelentně tolerován, ale tato dávka nedostačovala pro udržení stabilně nízkých hodnot D-dimerů a normální koncentrace hemoglobinu. Kombinace 10 mg lenalidomidu a 50 mg thalidomidu denně stabilizovala nemoc na dlouhých 9 měsíců.
Background: Multiple angiomatosis is a rare disease with angiomatous formations in multiple organs and tissues and associated with a risk of fatal bleeding. Case description: In this patient, the bones, pleural and peritoneal cavities and digestive tract were involved. The patient had long-term been administered zoledronate that provided relief from bone pain as early as after the second dose. The effect of antiangiogenics was evaluated on CT and MRI. Since angiomatous proliferation is associated with chronic disseminated intravascular coagulation (DIC) and anaemisation, blood count and fibrinogen as well as D-dimer and soluble fibrin monomer concentrations are also used to assess treatment response. Results: Before treatment, D-dimer levels were in excess of 20 μg/mL, fibrinogen 1.4 g/L and soluble fibrin monomers were at measurable levels. During treatment with interferon α at a dose of 6 million units 3 times a week with the dose reduction after 10 month, the median fibrinogen concentration increased to 1.5 (1.2–2.0) g/L, the median D-dimer levels declined to 17.2 (13.4–20.0) μg/mL and fibrin monomers were still detectable. Thalidomide therapy (100 mg/day) provided reduction in the median D-dimer levels to 6.07 (4.71–10.21) μg/ml and increase in median fibrinogen concentration to 1.9 g/L; soluble fibrin monomers were unidentifiable. CT imaging suggested significant reduction of angiomatous mass. Progressing neuropathy required dose reduction of thalidomide to 50 mg/day, leading to D-dimer increase. Lenalidomide 10 mg/day provided an increase in median D-dimer concentration to 10.8 (10.8–17.35) and decline in the level of haemoglobin to a median of 124 (135–117) g/L. Soluble fibrin monomers became detectable again. Therefore, a low dose of lenalidomide 10 mg/day was combined with thalidomide 100 mg and, subsequently, 50 mg/day. Treatment with lenalidomide 10 mg and thalidomide 50 mg provided median D-dimer levels of 9.32 and the disease has remained stable for 9 months. Conclusion: Thalidomide 100 mg/day stabilized multiple angiomatosis better than interferon alfa. Thalidomide 50 mg/day was insufficient to maintain disease stability. Lenalidomide at a dose of 10 mg was tolerated really well but this dose was insufficient to maintain low D-dimer levels and normal haemoglobin concentrations. The combination of lenalidomide 10 mg and thalidomide 50 mg daily stabilized the disease for 9 months.
- Klíčová slova
- hemangiomatosis, angiomatosis, D-dimery, chronická diseminovaná intravaskulární koagulace, PET-CT zobrazení,
- MeSH
- angiomatóza diagnóza farmakoterapie komplikace MeSH
- bisfosfonáty terapeutické užití MeSH
- břišní dutina patologie MeSH
- chronická nemoc MeSH
- diseminovaná intravaskulární koagulace diagnóza MeSH
- farmakologické účinky MeSH
- fibrin-fibrinogen - produkty degradace diagnostické užití MeSH
- financování organizované MeSH
- hemangiom diagnóza MeSH
- hemokoagulace MeSH
- interferon alfa škodlivé účinky terapeutické užití MeSH
- klinický obraz nemoci MeSH
- kombinovaná farmakoterapie MeSH
- krevní obraz MeSH
- lenalidomid MeSH
- lidé středního věku MeSH
- magnetická rezonanční tomografie MeSH
- pozitronová emisní tomografie MeSH
- příznaky a symptomy MeSH
- protinádorové látky MeSH
- retroperitoneální prostor patologie MeSH
- tenké střevo patologie MeSH
- thalidomid analogy a deriváty farmakologie terapeutické užití MeSH
- vyšetření krevní srážlivosti MeSH
- výsledek terapie MeSH
- žaludek patologie MeSH
- Check Tag
- lidé středního věku MeSH
- mužské pohlaví MeSH
- Publikační typ
- kazuistiky MeSH
19F magnetic resonance imaging (MRI) using fluoropolymer tracers has recently emerged as a promising, non-invasive diagnostic tool in modern medicine. However, despite its potential, 19F MRI remains overlooked and underused due to the limited availability or unfavorable properties of fluorinated tracers. Herein, we report a straightforward synthetic route to highly fluorinated 19F MRI nanotracers via aqueous dispersion polymerization-induced self-assembly of a water-soluble fluorinated monomer. A polyethylene glycol-based macromolecular chain-transfer agent was extended by RAFT-mediated N-(2,2,2-trifluoroethyl)acrylamide (TFEAM) polymerization in water, providing fluorine-rich self-assembled nanoparticles in a single step. The resulting nanoparticles had different morphologies and sizes ranging from 60 to 220 nm. After optimizing their structure to maximize the magnetic relaxation of the fluorinated core, we obtained a strong 19F NMR/MRI signal in an aqueous environment. Their non-toxicity was confirmed on primary human dermal fibroblasts. Moreover, we visualized the nanoparticles by 19F MRI, both in vitro (in aqueous phantoms) and in vivo (after subcutaneous injection in mice), thus confirming their biomedical potential.
Xanthophylls in light harvesting complexes perform a number of functions ranging from structural support to light-harvesting and photoprotection. In the major light harvesting complex of photosystem II in plants (LHCII), the innermost xanthophyll binding pockets are occupied by lutein molecules. The conservation of these sites within the LHC protein family suggests their importance in LHCII functionality. In the present work, we induced the photoprotective switch in LHCII isolated from the Arabidopsis mutant npq1lut2, where the lutein molecules are exchanged with violaxanthin. Despite the differences in the energetics of the pigments and the impairment of chlorophyll fluorescence quenching in vivo, we show that isolated complexes containing violaxanthin are still able to induce the quenching switch to a similar extent to wild type LHCII monomers. Moreover, the same spectroscopic changes take place, which suggest the involvement of the terminal emitter site (L1) in energy dissipation in both complexes. These results indicate the robust nature of the L1 xanthophyll binding domain in LHCII, where protein structural cues are the major determinant of the function of the bound carotenoid.
