Capping Reaction Research Articles

Structural and functional insights into the enzymatic plasticity of the SARS-CoV-2 NiRAN domain

The enzymatic activity of the SARS-CoV-2 nidovirus RdRp-associated nucleotidyltransferase (NiRAN) domain is essential for viral propagation, with three distinct activities associated with modification of the nsp9N terminus, NMPylation, RNAylation, and deRNAylation/capping via a GDP-polyribonucleotidyltransferase reaction. The latter two activities comprise an unconventional mechanism for initiating viral RNA 5' cap formation, while the role of NMPylation is unclear. The structural mechanisms for these diverse enzymatic activities have not been properly delineated. Here, we determine high-resolution cryoelectron microscopy (cryo-EM) structures of catalytic intermediates for the NMPylation and deRNAylation/capping reactions, revealing diverse nucleotide binding poses and divalent metal ion coordination sites to promote its repertoire of activities. The deRNAylation/capping structure explains why GDP is a preferred substrate for the capping reaction over GTP. Altogether, these findings enhance our understanding of the promiscuous coronaviral NiRAN domain, a therapeutic target, and provide an accurate structural platform for drug development.

Access to High-Purity 7m G-cap RNA in Substantial Quantities by a Convenient All-Chemical Solid-Phase Method.

Given the importance of mRNA with 5'-cap, easy access to RNA substrates with different 7m G caps, of high quality and in large quantities is essential to elucidate the roles of RNA and the regulation of underlying processes. In addition to existing synthetic routes to 5'-cap RNA based on enzymatic, chemical or chemo-enzymatic methods, we present here an all-chemical method for synthetic RNA capping. The novelty of this study lies in the fact that the capping reaction is performed on solid-support after automated RNA assembly using commercial 2'-O-propionyloxymethyl ribonucleoside phosphoramidites, which enable final RNA deprotection under mild conditions while preserving both 7m G-cap and RNA integrity. The capping reaction is efficiently carried out between a 5'-phosphoroimidazolide RNA anchored on the support and 7m GDP in DMF in the presence of zinc chloride. Substantial amounts of 7m G-cap RNA (from 1 to 28 nucleotides in length and of any sequence with or without internal methylations) containing various cap structures (7m GpppA, 7m GpppAm , 7m Gpppm6 A, 7m Gpppm6 Am , 7m GpppG, 7m GpppGm ) were obtained with high purity after IEX-HPLC purification. This capping method using solid-phase chemistry is convenient to perform and provides access to valuable RNA substrates as useful research tools to unravel specific issues regarding cap-related processes.

Structural basis and dynamics of Chikungunya alphavirus RNA capping by nsP1 capping pores

Alphaviruses are emerging positive-stranded RNA viruses which replicate and transcribe their genomes in membranous organelles formed in the cell cytoplasm. The nonstructural protein 1 (nsP1) is responsible for viral RNA capping and gates the replication organelles by assembling into monotopic membrane-associated dodecameric pores. The capping pathway is unique to Alphaviruses; beginning with the N7 methylation of a guanosine triphosphate (GTP) molecule, followed by the covalent linkage of an m7GMP group to a conserved histidine in nsP1 and the transfer of this cap structure to a diphosphate RNA. Here, we provide structural snapshots of different stages of the reaction pathway showing how nsP1 pores recognize the substrates of the methyl-transfer reaction, GTP and S-adenosyl methionine (SAM), how the enzyme reaches a metastable postmethylation state with SAH and m7GTP in the active site, and the subsequent covalent transfer of m7GMP to nsP1 triggered by the presence of RNA and postdecapping reaction conformational changes inducing the opening of the pore. In addition, we biochemically characterize the capping reaction, demonstrating specificity for the RNA substrate and the reversibility of the cap transfer resulting in decapping activity and the release of reaction intermediates. Our data identify the molecular determinants allowing each pathway transition, providing an explanation for the need for the SAM methyl donor all along the pathway and clues about the conformational rearrangements associated to the enzymatic activity of nsP1. Together, our results set ground for the structural and functional understanding of alphavirus RNA-capping and the design of antivirals.

Structure, function and dynamics of alphavirus capping pores.

Positive-sense single stranded RNA viruses such as coronaviruses, flaviviruses or alphaviruses, carry out transcription and replication inside virus-induced membranous organelles within host cells. Remodelling of the host cell membranes for formation of these organelles is coupled to the membrane association of the viral replication complexes and RNA synthesis. These viral niches allow for concentration of metabolites and proteins for the synthesis of viral RNA, also preventing its detection by the cellular innate immune system. I will present the cryo-EM structural characterization of the chikungunya virus non-structural protein 1, the viral protein responsible for RNA capping and membrane binding of the viral replication machinery, and the capping mechanism. The structures show the enzyme in its active form assembled in a monotopic membrane-associated dodecameric rings. It provides the structural basis for the coupling between membrane binding and allosteric activation of the capping enzyme. We also show different stages of the reaction pathway carried out by the enzyme, showing how nsP1 pores recognize the substrates of the methyl-transfer reaction, GTP and SAM, how it reaches a metastable post-methylation state with SAH and m7GTP in the active site, the subsequent covalent transfer of m7GMP to nsP1 and post-reaction conformational changes triggering the opening of the pore. In addition, we biochemically characterize the capping reaction, demonstrating specificity for the RNA substrate and the reversibility of the cap transfer resulting in decapping activity and the release of intermediates of the reaction. Our data identify the molecular determinants allowing each transition, provide explanation for the need for the SAM methyl donor all along the pathway and new clues about the conformational rearrangements associated to the enzymatic activity of nsP1. Together our results set new ground for the structural and functional understanding of alphavirus RNA-capping and the design of antivirals.

Synthesis of a Novel Bifunctional Epoxy Double-Decker Silsesquioxane: Improvement of the Thermal Stability and Dielectric Properties of Polybenzoxazine.

In this study a new type of bifunctional epoxy compound (DDSQ-EP) based on double-decker silsesquioxane (DDSQ) was synthesized by process of alkaline hydrolysis condensation of phenyltrimethoxysilane and corner capping reaction with dichloromethylvinylsilane, followed by epoxidation reaction of vinyl groups. The resultant structures were confirmed using Fourier transform infrared spectrometry, nuclear magnetic resonance spectrometry and time-of-flight mass spectrometry, respectively. The DDSQ-EP was incorporated into polybenzoxazine to obtain the PBZ/DDSQ-EP nanocomposites. The uniform dispersion of DDSQ-EP in the nanocomposites was verified by X-ray diffraction and scanning electron microscope. The reactions occurred during the curing of the composites and were investigated using infrared spectroscopy of segmented cures. Dynamic mechanical analysis and thermal gravimetric analysis indicated that the storage modulus, glass transition temperature and thermal stability of PBZ/DDSQ-EP were increased in comparison with pure benzoxazine resins. Assessment of dielectric properties demonstrated that the dielectric permittivity and dielectric loss of polybenzoxazine decreased slightly because of the addition of DDSQ-EP.

The mechanism of RNA capping by SARS-CoV-2

The RNA genome of SARS-CoV-2 contains a 5′ cap that facilitates the translation of viral proteins, protection from exonucleases and evasion of the host immune response1–4. How this cap is made in SARS-CoV-2 is not completely understood. Here we reconstitute the N7- and 2′-O-methylated SARS-CoV-2 RNA cap (7MeGpppA2′-O-Me) using virally encoded non-structural proteins (nsps). We show that the kinase-like nidovirus RdRp-associated nucleotidyltransferase (NiRAN) domain5 of nsp12 transfers the RNA to the amino terminus of nsp9, forming a covalent RNA–protein intermediate (a process termed RNAylation). Subsequently, the NiRAN domain transfers the RNA to GDP, forming the core cap structure GpppA-RNA. The nsp146 and nsp167 methyltransferases then add methyl groups to form functional cap structures. Structural analyses of the replication–transcription complex bound to nsp9 identified key interactions that mediate the capping reaction. Furthermore, we demonstrate in a reverse genetics system8 that the N terminus of nsp9 and the kinase-like active-site residues in the NiRAN domain are required for successful SARS-CoV-2 replication. Collectively, our results reveal an unconventional mechanism by which SARS-CoV-2 caps its RNA genome, thus exposing a new target in the development of antivirals to treat COVID-19.

Complete Chemical Synthesis of Minimal Messenger RNA by Efficient Chemical Capping Reaction.

Site-specific chemical modification of mRNA can improve its translational efficiency and stability. For this purpose, it is desirable to develop a complete chemical synthesis method for chemically modified mRNA. The key is a chemical reaction that introduces a cap structure into the chemically synthesized RNA. In this study, we developed a fast and quantitative chemical capping reaction between 5'-phosphorylated RNA and N7-methylated GDP imidazolide in the presence of 1-methylimidazole in the organic solvent dimethyl sulfoxide. It enabled quantitative preparation of capping RNA within 3 h. We prepared chemically modified 107-nucleotide mRNAs, including N6-methyladenosine, insertion of non-nucleotide linkers, and 2'-O-methylated nucleotides at the 5' end and evaluated their effects on translational activity in cultured HeLa cells. The results showed that mRNAs with non-nucleotide linkers in the untranslated regions were sufficiently tolerant to translation and that mRNAs with the Cap_2 structure had higher translational activity than those with the Cap_0 structure.

Effect of PbSO4-Oleate Coverage on Cesium Lead Halide Perovskite Quantum Dots to Control Halide Exchange Kinetics.

The selective control of halide ion exchange in metal halide perovskite quantum dots (PQDs) plays an important role in determining their band gap and composition. In this study, CsPbX3 (X = Cl−, Br−, and I−) PQDs were self-assembled with PbSO4-oleate to form a peapod-like morphology to selectively control halide ion exchange. Considering the distinct absorption and bright luminescence characteristics of these PQDs, in situ UV-Vis. absorption and fluorescence spectroscopies were employed to monitor the time-dependent band gap and compositional changes of the PQDs. We determined that the halide exchange in the capped PQDs is hindered—unlike the rapid anion exchange in noncapped PQDs—by a reduction in the halide exchange kinetic rate depending on the extent of coverage of the PQDs. Thus, we tracked the halide ion exchange kinetics between CsPbBr3 and CsPbI3 PQDs, depending on the coverage, using in situ UV-Vis. absorption/photoluminescence spectroscopy. We regulated the halide exchange reaction rate by varying the capping reaction temperature of the PQDs. The capping hindered the halide exchange kinetics and increased the activation energy. These results will enable the development of white LEDs, photovoltaic cells, and photocatalysts with alternative structural designs based on the divalent composition of CsPbX3 PQDs.

Crack- and Shrinkage-Free Ethylene-Bridged Polysilsesquioxane Film Prepared by a Hydrosilylation Reaction.

With the aim of developing an improved strategy for the preparation of ethylene-bridged polysilsesquioxanes as thermal insulator materials, this paper describes the synthesis of a crack- and shrinkage-free ethylene-bridged polysilsesquioxane film by the hydrosilylation reaction of hydrodimethyl-silylated oligomethylsilsesquioxane (MSQ-SiH) and dimethylvinyl-silylated oligomethylsilsesquioxane (MSQ-SiVi) in the presence of Karstedt’s catalyst. Polysilsesquioxane precursors were prepared by the sol–gel reaction of triethoxymethylsilane and the successive capping reaction with chlorodimethylsilane and chlorodimethylvinylsilane. The obtained ethylene-bridged polysilsesquioxane film showed lower density and thermal diffusivity (1.13 g/cm3 and 1.15 × 10–7 m2/s, respectively) than a polymethylsilsesquioxane film (1.34 g/cm3 and 1.36 × 10–7 m2/s, respectively). As a result of the introduction of the SiCCSi ethylene bridge, the thermal insulation property of the polysilsesquioxane film was enhanced.

Synthesis of Core@Shell Cu‐Ni@Pt‐Cu Nano‐Octahedra and Their Improved MOR Activity

AbstractFabrication of 3d metal‐based core@shell nanocatalysts with engineered Pt‐surfaces provides an effective approach for improving the catalytic performance. The challenges in such preparation include shape control of the 3d metallic cores and thickness control of the Pt‐based shells. Herein, we report a colloidal seed‐mediated method to prepare octahedral CuNi@Pt‐Cu core@shell nanocrystals using CuNi octahedral cores as the template. By precisely controlling the synthesis conditions including the deposition rate and diffusion rate of the shell‐formation through tuning the capping ligand, reaction temperature, and heating rate, uniform Pt‐based shells can be achieved with a thickness of <1 nm. The resultant carbon‐supported CuNi@Pt‐Cu core@shell nano‐octahedra showed superior activity in electrochemical methanol oxidation reaction (MOR) compared with the commercial Pt/C catalysts and carbon‐supported CuNi@Pt‐Cu nano‐polyhedron counterparts.

Synthesis of Core@Shell Cu-Ni@Pt-Cu Nano-Octahedra and Their Improved MOR Activity.

Fabrication of 3d metal-based core@shell nanocatalysts with engineered Pt-surfaces provides an effective approach for improving the catalytic performance. The challenges in such preparation include shape control of the 3d metallic cores and thickness control of the Pt-based shells. Herein, we report a colloidal seed-mediated method to prepare octahedral CuNi@Pt-Cu core@shell nanocrystals using CuNi octahedral cores as the template. By precisely controlling the synthesis conditions including the deposition rate and diffusion rate of the shell-formation through tuning the capping ligand, reaction temperature, and heating rate, uniform Pt-based shells can be achieved with a thickness of <1 nm. The resultant carbon-supported CuNi@Pt-Cu core@shell nano-octahedra showed superior activity in electrochemical methanol oxidation reaction (MOR) compared with the commercial Pt/C catalysts and carbon-supported CuNi@Pt-Cu nano-polyhedron counterparts.

Fabrication of a nonionic self-emulsifiable waterborne epoxy curing agent with high cure properties

Waterborne epoxy curing agent plays a key part in the waterborne epoxy resin coatings, directly affecting the physical and chemical properties of the coating film. Here, we present triethylene tetramine (TETA), polypropylene glycol diglycidyl ether (PPGDGE), epoxy resin E20 and phenyl glycidyl ether (PGE) as reactants to fabricate a novel nonionic self-emulsifiable waterborne epoxy curing agent for oily epoxy resin through three steps of chain extension reaction, addition reaction, and capping reaction. The curing agent achieved excellent emulsifying and curing properties under the condition that TETA reacted with PPGDGE at 65°C for 4 h with the molar ratio of 2.2:1, 40% primary amine was added with E20 at 65°C for 3 h and 125% residual primary amine was capped with PGE at 65°C for 3 h. The as-prepared waterborne epoxy curing agent was characterized by Fourier transform-infrared spectroscopy and surface tension. The cured film of emulsified oily epoxy resin (E51) prepared by the as-prepared curing agent indirectly was characterized by scanning electron microscopy, thermogravimetric analysis and differential scanning calorimetry tests. The 2K waterborne epoxy coating film prepared by the as-prepared nonionic self-emulsifiable curing agent and E51 displayed better thermal property, surface appearance, flexibility, impact resistance, adhesion, and chemical corrosion resistance.

Mutations on VEEV nsP1 relate RNA capping efficiency to ribavirin susceptibility

Alphaviruses are arthropod-borne viruses of public health concern. To date no efficient vaccine nor antivirals are available for safe human use. During viral replication the nonstructural protein 1 (nsP1) catalyzes capping of genomic and subgenomic RNAs. The capping reaction is unique to the Alphavirus genus. The whole three-step process follows a particular order: (i) transfer of a methyl group from S-adenosyl methionine (SAM) onto a GTP forming m7GTP; (ii) guanylylation of the enzyme to form a m7GMP-nsP1adduct; (iii) transfer of m7GMP onto 5′-diphosphate RNA to yield capped RNA. Specificities of these reactions designate nsP1 as a promising target for antiviral drug development. In the current study we performed a mutational analysis on two nsP1 positions associated with Sindbis virus (SINV) ribavirin resistance in the Venezuelan equine encephalitis virus (VEEV) context through reverse genetics correlated to enzyme assays using purified recombinant VEEV nsP1 proteins. The results demonstrate that the targeted positions are strongly associated to the regulation of the capping reaction by increasing the affinity between GTP and nsP1. Data also show that in VEEV the S21A substitution, naturally occurring in Chikungunya virus (CHIKV), is a hallmark of ribavirin susceptibility. These findings uncover the specific mechanistic contributions of these residues to nsp1-mediated methyl-transfer and guanylylation reactions.

Synthesis of3,13‐diglycidyloxypropyloctaphenyldouble‐decker polyhedral oligomeric silsesquioxane and the thermal reaction properties with thermosetting phenol‐formaldehyde resin

Abstract3,13‐Diglycidyloxypropyloctaphenyl double‐decker silsesquioxane (EP‐DDSQ) was synthesized by process of alkaline hydrolysis condensation of phenyltrimethoxysilane and corner capping reaction of methyldichlorosilane, followed by hydrosilylation with allyl glycidyl ether, and the resultant structure was confirmed by fourier transform infrared spectrometer (FTIR) and nuclear magnetic resonance (NMR), respectively. The thermosetting phenol‐formaldehyde (PF) resin was then modified by EP‐DDSQ, and the reactivity of PF resin with EP‐DDSQ and thermal pyrolysis of modified cured resin were investigated by differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). The surface morphologies of modified resins at high temperature were characterized with field emission scanning electron microscope (FESEM), and chemical structure of modified resins was analyzed through X‐ray photoelectron spectrometer (XPS). The results showed that the appropriate addition of EP‐DDSQ did not affect the curing temperature of the PF resin itself, but could improve the heat resistance of the system. When the amount of EP‐DDSQ added was 10%, the initial degradation temperature of PF resin was increased by 49.31°C, and when the amount of EP‐DDSQ added was 16%, the char yield of which was reached up to 61.39%, compared with that of pure PF resin (TGA1,000°Cof 57.62%) at Ar atmosphere. More importantly, the modified resin formed a regular and dense layer of SiC and SiOxceramic on the surface after ablation in the muffle furnace at 800°C air atmosphere, which is very important for ablative resistant materials.

Custom-Tailoring Loose Nanofiltration Membrane for Precise Biomolecule Fractionation: New Insight into Post-Treatment Mechanisms.

Loose nanofiltration (NF) membranes with diverse selectivity can meet the great demands in various bioseparation applications. Thus, a facile strategy to tune the properties such as pore size, surface charge, and hydrophilicity of the NF membrane is required to produce tailor-made loose NF membranes without changing the existing production line. Herein, we systematically investigated the post-treatment of the nascent poly(piperazine amide) NF membranes using different reagents (organic acids, weak bases, organic solvents and ionic liquid (IL)). Various characterizations revealed that the skin/separation layer became looser and permeance was promoted with the decrease of salt rejection in varying degrees. It was found that the O/N ratio did not rigorously represent the cross-linking degree of the skin layer, because besides the hydrolysis of the residual acyl chloride impeding the amido bond formation, the breaking of existing amido bonds and the grafting of free trimesoyl chloride molecules on the nascent membranes could also increase the O/N ratio during post-treatments. Then three mechanisms including hydrolysis, swelling rearrangement and capping reaction effects were proposed to better understand the membrane properties variations. All these effects resulted in larger pore size of the NF membrane, and the hydrolysis/capping effect might increase negative charge and hydrophilicity on the membrane, while the swelling rearrangement could produce less defective skin structure. These three effects might be involved together during a single treatment. Finally, the NF membrane post-treated by N-hexane could efficiently separate antibiotics and NaCl with the highest permeate flux, whereas the one post-treated by ionic liquid outperformed others for the decoloration of cane molasses (much more efficient than NF270, DL, and NTR7450 membranes). The long-term operating stability of the post-treated membranes selected was also confirmed by a continuous crossflow filtration for 15 h with regular alkaline cleaning.

Development of an RNA Expression Platform Controlled by Viral Internal Ribosome Entry Sites.

Since 1990, many nucleic acid expression platforms consisting of DNA or RNA have been developed. However, although RNA expression platforms have been relatively neglected, several such platforms capped at the 5' end of RNA by an anti-reverse cap analog have now been developed. At the same time, the capping reaction is a bottleneck in the production of such platforms, with high cost and low efficiency. Here, we investigated several viral and eukaryotic internal ribosome entry sites (IRESs) to develop an optimal RNA expression platform, because IRES-dependent translation does not require a capping step. RNA expression platforms constructed with IRESs from the 5' untranslated regions of the encephalomyocarditis virus (EMCV) and the intergenic region of the cricket paralysis virus (CrPV) showed sufficient expression efficiency compared with cap-dependent RNA expression platforms. However, eukaryotic IRESs exhibited a lower viral IRES expression efficiency. Interestingly, the addition of a poly(A) sequence to the 5' end of the coxsackievirus B3 (CVB3) IRES (pMA-CVB3) increased the expression level compared with the CVB3 IRES without poly(A) (pCVB3). Therefore, we developed two multiexpression platforms (termed pMA-CVB3-EMCV and pCrPV-EMCV) by combining the IRESs of CVB3, CrPV, and EMCV in a single-RNA backbone. The pMA-CVB3-EMCV-derived RNA platform showed the highest expression level. Moreover, it clearly exhibited expression in mouse muscles in vivo. These RNA expression platforms prepared using viral IRESs will be useful in developing potential RNA-based prophylactic or therapeutic vaccines, because they have better expression efficiency and do not need a capping step.

Effect of Mono-Substituents in Heptaisobutyl-Substituted Polyhedral Octasilsesquioxanes on Orientationally Disordered Phase Transition

Abstract A series of mono-functionalized polyhedral octasilsesquioxane (POSS) derivatives (R-POSSs) were prepared by corner capping reaction of heptaisobutyl incompletely condensed POSS and their orientationally disordered (OD) phase transition properties studied by DSC, XRD, and solid state NMR analyses. According to DSC analysis, octaisobutyl-POSS (iBu-POSS) showed the highest OD phase transition temperature among R-POSSs, whereas the OD phase transition temperatures decreased with replacing the mono-substituents smaller and larger than the isobutyl unit. The solid state NMR analysis suggests that the OD phase transition was caused by increasing the mobility of the substituent not by rotating the POSS cage. The composite films were prepared by drop-casting of a chloroform solution of PMMA with heptaisobutyl-butyl-POSS (Bu-POSS). No appearance change of films was observed despite the heating and cooling of the films on the hotplate due to the absence of drastic density difference below and above the OD phase transition.

The effect of amine-functionalized 1,1-diphenylethylene (DPE) derivatives on end-capping reactions and the simulation of their precision for sequence control

Three amine-functionalized 1,1-diphenylethylene (DPE) derivatives (DPE-(NMe2)2, DPE-NMe2 and DPE-SiH/NMe2) were used to study the end-capping reactions of poly(styryl)lithium (PS-Li) for investigating the effect on end-capping of electron-donating groups in DPE structures. In situ1H NMR was used to measure the kinetic parameters of end-capping reactions. When the DPE derivative was held constant in the PS-Li capping reaction, the kinetic parameter (kSD) did not change with the variance of either concentration or degree of polymerization of PS-Li. Additionally, the kSD value changed dramatically when various electronic properties of the DPE derivative substituents were modified. Electron-withdrawing groups accelerated the end-capping reaction, while electron-donating groups decelerated it. The kSD experimental values of DPE-(NMe2)2, DPE-NMe2 and DPE-SiH/NMe2 were found to be 0.155 × 10−3, 1.26 × 10−3, and 6.24 × 10−3 L1/2 mol−1/2 s−1, respectively. An intuitive model was established to simulate the precision of the sequence-controlled copolymerization with DPE derivatives based on investigations of end-capping kinetics. The model depicts the degree of fictitious styrene propagation for the chain insertion of one DPE derivative unit. The fictitious degrees of polymerization of styrene (FDPs) for DPE-NMe2 and DPE-SiH/NMe2 were calculated to be 37 and 13, respectively, suggesting that using DPE derivatives containing an electron-withdrawing group in the synthesis of sequence-controlled polymers might lead to a more precise structure.

Synthesis and characterization of well-defined poly(2-deoxy-2-methacrylamido-d-glucose) and its biopotential block copolymers via RAFT and ROP polymerization

Well-defined homopolymers of 2-deoxy-2-methacrylamido-d-glucose (MAG) were synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization using 4-cyanopentanoic acid-4-dithiobenzoate as chain transfer agent (CTA) and 2,2′-azobisisobutyronitrile (AIBN) as initiator. The effect of polymerization conditions such as molar ratio of RAFT agent to initiator and monomer to RAFT agent on conversion and molecular mass characteristics of prepared polymers was estimated. Kinetics of the polymerization under various reaction conditions was studied as a function of time and conversion using in situ1H NMR spectroscopy. The living character of polymerization was demonstrated by carriyng out the further chain extension experiments using isolated PMAG-CTA as a macroRAFT agent. For that, the PMAG with higher molecular weight, as well as block copolymer with acrylic acid were synthesized and characterized using 1H NMR and SEC analysis. A novel amphiphilic copolymer representing a hybrid of glycopolymer and polypeptide fragments was synthesized using RAFT and ring-opening polymerization (ROP). In this case, dithiobenzoate end-group of PMAG was transformed via simultaneous reduction and in situ thiol capping reaction by cysteamine hydrochloride into amino functionality suitable for polymerization of N-carboxyanhydrides (NCA). Two block copolymers of MAG with poly-l-phenylalanine (PPhe) differed with hydrophobic block length were synthesized, characterized and used for preparation of particles via copolymer self-assembly. The hydrodynamic diameter, morphology and cytotoxicity of polymer particles based on PMAG-b-PPhe were evaluated using DLS, TEM and MTT-assay, respectively. Moreover, the encapsulation of model compound (fluorescent dye rhodamine 6G) inside PMAG-b-PPhe micelles, as well as its release, were also tested.

Synthesis and properties of hyperbranched polymers by polymerization of an AB3-type incompletely condensed cage silsesquioxane (IC-POSS) monomer

An incompletely condensed cage silsesquioxane (IC-POSS)-based AB3-type monomer, tris(dimethylsilyl)-p-vinylhexaisobutyl-IC-POSS (3), was prepared by the corner-cleaved reaction of vinylheptaisobutyl-POSS (1) and subsequent capping reaction with chlorodimethylsilane. The structure of 3 was confirmed by FT-IR; 1H-, 13C-, and 29Si-NMR; and HR-FAB-MS analyses. These analyses also suggested 12.5 mol% contamination of tris(dimethylsilyl)-heptaisobutyl-IC-POSS (4). The hydrosilylation polymerization of 3 as an AB3-type IC-POSS monomer in the presence of Karstedt’s catalyst in toluene at 80 °C overnight provided a soluble polymer with a number average molecular weight (Mn) and molecular weight distribution (Mw/Mn) of the polymer that are 3.9 × 103 and 1.39, respectively. The resulting hyperbranched polymer (HB) reacted with two molar equivalents of isoprenol or allylheptaisobutyl-POSS against the Si–H groups in the presence of Karstedt’s catalyst in toluene at 80 °C for 7 h. The HB polymers before and after post functionalization with isoprenol or allylheptaisobutyl-POSS showed 1% weight loss at 298, 285, and 358 °C, respectively, under N2. Introducing POSS units at the terminus significantly increased the thermal stability. The XRD pattern of the HB polymers showed a denser structure formed in the allylheptaisobutyl-POSS-terminated HB polymer. The refractive index of the HB polymer before post functionalization was 1.4376. Post functionalization with isoprenol and allylheptaisobutyl-POSS increased the refractive index to 1.4464 and 1.4573, respectively. An incompletely condensed cage silsesquioxane (IC-POSS)-based AB3-type monomer, tris(dimethylsilyl)-p-vinylhexaisobutyl-IC-POSS (3), was prepared by the corner-cleaved reaction of vinylheptaisobutyl-POSS (1) and subsequent capping reaction with chlorodimethylsilane. The hydrosilylation polymerization of 3 in the presence of Karstedt’s catalyst in toluene provided a soluble polymer. The refractive index of the unmodified polymer film was 1.4376. Post functionalization with isoprenol increased the dispersibility in polar organic solvents and significantly increased the glass-transition temperature. Post functionalization with allylheptaisobutyl-POSS significantly increased the thermal stability. Their refractive index varied from 1.4464 to 1.4573 upon post functionalization.