Dynamics of end-grafted polystyrene brushes in theta solvents

The dynamics of the concentration fluctuations in end-grafted polystyrene brushes in a theta solvent (cyclohexane) are probed by evanescent wave dynamic light scattering at different wavevectors q and temperatures. When the solvent quality changes from marginal to poor, the relaxation function C(q, t) exhibits strong effects as compared with the smooth variation of the brush density profile. From a single exponential above 50 °C, C(q, t) becomes a two-step decay function. The fast decay is still assigned to the cooperative diffusion albeit slower than in the good solvent regime whereas the slow nonexponential and nondiffusive process might relate to microsegragated and/or chain dynamics in the present polydisperse brush. The relaxation function of the present three brushes with different grafting density reveals similarities and disparities between wet brushes and semidilute polymer solutions. © 2006 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 44: 3590–3597, 2006     

Synthesis and characterization of novel pH-responsive microgels based on tertiary amine methacrylates

Emulsion polymerization of 2-(diethylamino)ethyl methacrylate (DEA) in the presence of a bifunctional cross-linker at pH 8-9 afforded novel pH-responsive microgels of 250-700 nm diameter. Both batch and semicontinuous syntheses were explored using thermal and redox initiators. Various strategies were evaluated for achieving colloidal stability, including charge stabilization, surfactant stabilization, and steric stabilization. The latter proved to be the most convenient and effective, and three types of well-defined reactive macromonomers were examined, namely, monomethoxy-capped poly(ethylene glycol) methacrylate (PEGMA), styrene-capped poly[2-(dimethylamino)ethyl methacrylate] (PDMA50-St), and partially quaternized styrene-capped poly[2-(dimethylamino)ethyl methacrylate] (10qPDMA50-St). The resulting microgels were pH-responsive, as expected. Dynamic light scattering and 1H NMR studies confirmed that reversible swelling occurred at low pH due to protonation of the tertiary amine groups on the DEA residues. The critical pH for this latex-to-microgel transition was around pH 6.5-7.0, which corresponds approximately to the known pKa of 7.0-7.3 for linear PDEA homopolymer. The microgel particles were further characterized by electron microscopy and aqueous electrophoresis studies. Their swelling and deswelling kinetics were investigated by turbidimetry. The PDEA-based microgels were compared to poly[2-(diisopropylamino)ethyl methacrylate] (PDPA) microgels prepared with identical macromonomer stabilizers. These PDPA-based microgels had a lower critical swelling pH of around pH 5.0-5.5, which correlates with the lower pKa of PDPA homopolymer. In addition, the kinetics of swelling for the PDPA microgels was somewhat slower than that observed for PDEA microgels; presumably this is related to the greater hydrophobic character of the former particles.     

Engineered SAM Synthetases for Enzymatic Generation of AdoMet Analogs with Photocaging Groups and Reversible DNA Modification in Cascade Reactions

Methylation and demethylation of DNA, RNA and proteins has emerged as a major regulatory mechanism. Studying the function of these modifications would benefit from tools for their site‐specific inhibition and timed removal. S‐Adenosyl‐L‐methionine (AdoMet) analogs in combination with methyltransferases (MTases) have proven useful to map or block and release MTase target sites, however their enzymatic generation has been limited to aliphatic groups at the sulfur atom. We engineered a SAM synthetase from Cryptosporidium hominis (PC‐ChMAT) for efficient generation of AdoMet analogs with photocaging groups that are not accepted by any WT MAT reported to date. The crystal structure of PC‐ChMAT at 1.87 Å revealed how the photocaged AdoMet analog is accommodated and guided engineering of a thermostable MAT from Methanocaldococcus jannaschii. PC‐MATs were compatible with DNA‐ and RNA‐MTases, enabling sequence‐specific modification (“writing”) of plasmid DNA and light‐triggered removal (“erasing”).     

Observation of slow down of polystyrene nanogels diffusivities in contact with swollen polystyrene brushes

The diffusion of dilute colloids in contact with swollen polymer brushes has been studied by evanescent wave dynamic light scattering. Two polystyrene nanogels with 16 nm and 42 nm radius were put into contact with three polystyrene brushes with varying grafting densities. Partial penetration of the nanogels within the brushes was revealed by the evanescent wave penetration depth-dependent scattering intensities. The experimental short-time diffusion coefficients of the penetrating particles were measured and found to strongly slow down as the nanoparticles get deeper into the brushes. The slow down is much more marked for the smaller (16 nm) nanogels, suggesting a size exclusion type of mechanism and the existence of a characteristic length scale present in the outer part of the brush.     

Synthesis and Conformational Analysis of Fluorinated Uridine Analogues Provide Insight into a Neighbouring-Group Participation Mechanism

Fluorinated nucleoside analogues have attracted much attention as anticancer and antiviral agents and as probes for enzymatic function. However, the lack of direct synthetic methods, especially for 2′,3′-dideoxy-2′,3′-difluoro nucleosides, hamper their practical utility. In order to design more efficient synthetic methods, a better understanding of the conformation and mechanism of formation of these molecules is important. Herein, we report the synthesis and conformational analysis of a 2′,3′-dideoxy-2′,3′-difluoro and a 2′-deoxy-2′-fluoro uridine derivative and provide an insight into the reaction mechanism. We suggest that the transformation most likely diverges from the S_N1 or S_N2 pathway, but instead operates via a neighbouring-group participation mechanism.     

Engineering of Therapeutic and Detoxifying Enzymes

Therapeutic enzymes present excellent opportunities for the treatment of human disease, modulation of metabolic pathways and system detoxification. However, current use of enzyme therapy in the clinic is limited as naturally occurring enzymes are seldom optimal for such applications and require substantial improvement by protein engineering. Engineering strategies such as design and directed evolution that have been successfully implemented for industrial biocatalysis can significantly advance the field of therapeutic enzymes, leading to biocatalysts with new-to-nature therapeutic activities, high selectivity, and suitability for medical applications. This minireview highlights case studies of how state-of-the-art and emerging methods in protein engineering are explored for the generation of therapeutic enzymes and discusses gaps and future opportunities in the field of enzyme therapy.     

Brownian diffusion close to a polymer brush

In an effort to control particle diffusion near surfaces, we have studied the dynamics of colloidal hard spheres and soft compliant star copolymers on surfaces coated with polymer brushes using evanescent wave dynamic light scattering. The same experiments provide information on the brush structure and confined particle motion. The penetration into dense polydisperse brushes is size- and solvent-dependent.