Aqueous solutions of some polymers exhibit a lower critical solution temperature (LCST); that is, they form phase-separated aggregates when heated above a threshold temperature. Such polymers found many promising (bio)medical applications, including in situ thermogelling with controlled drug release, polymer-supported radiotherapy (brachytherapy), immunotherapy, and wound dressing, among others. Yet, despite the extensive research on medicinal applications of thermoresponsive polymers, their biodistribution and fate after administration remained unknown. Thus, herein, they studied the pharmacokinetics of four different thermoresponsive polyacrylamides after intramuscular administration in mice. In vivo, these thermoresponsive polymers formed depots that subsequently dissolved with a two-phase kinetics (depot maturation, slow redissolution) with half-lives 2 weeks to 5 months, as depot vitrification prolonged their half-lives. Additionally, the decrease of TCP of a polymer solution increased the density of the intramuscular depot. Moreover, they detected secondary polymer depots in the kidneys and liver; these secondary depots also followed two-phase kinetics (depot maturation and slow dissolution), with half-lives 8 to 38 days (kidneys) and 15 to 22 days (liver). Overall, these findings may be used to tailor the properties of thermoresponsive polymers to meet the demands of their medicinal applications. Their methods may become a benchmark for future studies of polymer biodistribution.

• Klíčová slova
• LCST, biodistribution, poly(2,2-difluoroethyl)acrylamide, poly(N,N-diethylacrylamide), poly(N-acryloylpyrolidine), poly(N-isopropylacrylamide), polyacrylamide, rational polymer design,
• MeSH
• myši MeSH
• polymery * MeSH
• teplota MeSH
• tkáňová distribuce MeSH
• uvolňování léčiv MeSH
• voda * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• polymery * MeSH
• voda * MeSH

Nucleolin is a multifunctional RNA Binding Protein (RBP) with diverse subcellular localizations, including the nucleolus in all eukaryotic cells, the plasma membrane in tumor cells, and the axon in neurons. Here we show that the glycine arginine rich (GAR) domain of nucleolin drives subcellular localization via protein-protein interactions with a kinesin light chain. In addition, GAR sequences mediate plasma membrane interactions of nucleolin. Both these modalities are in addition to the already reported involvement of the GAR domain in liquid-liquid phase separation in the nucleolus. Nucleolin transport to axons requires the GAR domain, and heterozygous GAR deletion mice reveal reduced axonal localization of nucleolin cargo mRNAs and enhanced sensory neuron growth. Thus, the GAR domain governs axonal transport of a growth controlling RNA-RBP complex in neurons, and is a versatile localization determinant for different subcellular compartments. Localization determination by GAR domains may explain why GAR mutants in diverse RBPs are associated with neurodegenerative disease.

• Klíčová slova
• axonal transport, cell size regulation, local translation, protein-membrane interaction, subcellular localization,
• MeSH
• axonální transport genetika   MeSH
• buněčné jadérko metabolismus   ultrastruktura   MeSH
• exprese genu MeSH
• fosfoproteiny chemie   genetika   metabolismus   MeSH
• HEK293 buňky MeSH
• HeLa buňky MeSH
• kineziny genetika   metabolismus   MeSH
• lidé MeSH
• messenger RNA genetika   metabolismus   MeSH
• mutace MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nervus ischiadicus cytologie   metabolismus   MeSH
• neurony cytologie   metabolismus   MeSH
• nukleolin MeSH
• primární buněčná kultura MeSH
• proteinové domény MeSH
• proteiny vázající RNA chemie   genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• spinální ganglia cytologie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• fosfoproteiny MeSH
• kineziny MeSH
• Kns2 protein, mouse MeSH Prohlížeč
• messenger RNA MeSH
• proteiny vázající RNA MeSH

The detailed functional mechanism of recoverin, which acts as a myristoyl switch at the rod outer-segment disk membrane, is elucidated by direct and replica-exchange molecular dynamics. In accord with NMR structural evidence and calcium binding assays, simulations point to the key role of enhanced calcium binding to the EF3 loop of the semiopen state of recoverin as compared to the closed state. This 2-4-order decrease in calcium dissociation constant stabilizes the semiopen state in response to the increase of cytosolic calcium concentration in the vicinity of recoverin. A second calcium ion then binds to the EF2 loop and, consequently, the structure of the protein changes from the semiopen to the open state. The latter has the myristoyl chain extruded to the cytosol, ready to act as a membrane anchor of recoverin.

• MeSH
• komplexní sloučeniny chemie   metabolismus   MeSH
• konformace proteinů MeSH
• magnetická rezonanční spektroskopie MeSH
• motivy EF-ruky MeSH
• mutace MeSH
• rekoverin chemie   genetika   metabolismus   MeSH
• simulace molekulární dynamiky MeSH
• skot MeSH
• termodynamika MeSH
• vápník chemie   metabolismus   MeSH
• vazba proteinů MeSH
• změna skupenství MeSH
• zvířata MeSH
• Check Tag
• skot MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• komplexní sloučeniny MeSH
• rekoverin MeSH
• vápník MeSH

Recoverin is a neuronal calcium sensor involved in vision adaptation that reversibly associates with cellular membranes via its calcium-activated myristoyl switch. While experimental evidence shows that the myristoyl group significantly enhances membrane affinity of this protein, molecular details of the binding process are still under debate. Here, we present results of extensive molecular dynamics simulations of recoverin in the proximity of a phospholipid bilayer. We capture multiple events of spontaneous membrane insertion of the myristoyl moiety and confirm its critical role in the membrane binding. Moreover, we observe that the binding strongly depends on the conformation of the N-terminal domain. We propose that a suitable conformation of the N-terminal domain can be stabilized by the disordered C-terminal segment or by binding of the target enzyme, i.e., rhodopsin kinase. Finally, we find that the presence of negatively charged lipids in the bilayer stabilizes a physiologically functional orientation of the membrane-bound recoverin.

• Publikační typ
• časopisecké články MeSH

Chemical diversity: Two SAM-dependent N-methyltransferases-LmbJ from the biosynthesis of the antibiotic lincomycin and CcbJ from celesticetin biosynthesis-have been characterized and compared. Both tested enzymes form multimers and are able to utilize N-demethyllincomycin, the natural substrate of LmbJ, with comparable efficiency.

• Klíčová slova
• N-methyltransferases, Streptomyces, antibiotics, lincomycin, natural products,
• MeSH
• antibakteriální látky biosyntéza   chemie   MeSH
• biokatalýza * MeSH
• linkomycin biosyntéza   chemie   MeSH
• linkosamidy biosyntéza   chemie   MeSH
• methyltransferasy chemie   metabolismus   MeSH
• molekulární konformace MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibakteriální látky MeSH
• celesticetin A MeSH Prohlížeč
• linkomycin MeSH
• linkosamidy MeSH
• methyltransferasy MeSH