Phosphodiester linkages, including those that join the nucleotides of DNA, are highly resistant to spontaneous hydrolysis. The rate of water attack at the phosphorus atom of phosphodiesters is known only as an upper limit, based on the hydrolysis of the dimethyl phosphate anion. That reaction was found to proceed at least 99% by C-O cleavage, at a rate suggesting an upper limit of 10 ؊15 s ؊1 for P-O cleavage of phosphodiester anions at 25°C. To evaluate the rate enhancement produced by P-O cleaving phosphodiesterases such as staphylococcal nuclease, we decided to establish the actual value of the rate constant for P-O cleavage of a simple phosphodiester anion. In dineopentyl phosphate, C-O cleavage is sterically precluded so that hydrolysis occurs only by P-O cleavage. Measurements at elevated temperatures indicate that the dineopentyl phosphate anion undergoes hydrolysis in water with a t 1/2 of 30,000,000 years at 25°C, furnishing an indication of the resistance of the internucleotide linkages of DNA to water attack at phosphorus. These results imply that staphylococcal nuclease (k cat ‫؍‬ 95 s ؊1 ) enhances the rate of phosphodiester hydrolysis by a factor of Ϸ10 17 . In alkaline solution, thymidylyl-3 -5 -thymidine (TpT) has been reported to decompose 10 5 -fold more rapidly than does dineopentyl phosphate. We find however that TpT and thymidine decompose at similar rates and with similar activation parameters, to a similar set of products, at pH 7 and in 1 M KOH. We infer that the decomposition of TpT is initiated by the breakdown of thymidine, not by phosphodiester hydrolysis.DNA hydrolysis ͉ DNA stability ͉ nuclease ͉ rate enhancement ͉ phosphate ester P hosphoric acid diesters are, in general, exceedingly unreactive in water (1-3), so that the phosphodiester linkages that join the nucleotides of DNA are highly resistant to spontaneous hydrolysis. By extrapolation of earlier model experiments at elevated temperatures, the uncatalyzed hydrolysis of dimethyl phosphate in neutral solution was found to proceed with an estimated rate constant of Ϸ2 ϫ 10 Ϫ13 s Ϫ1 at 25°C, corresponding to a half-time of 140,000 years. That reaction was found to proceed at least 99% by C-O cleavage, suggesting an upper limit of Ϸ1 ϫ 10 Ϫ15 s Ϫ1 at 25°C on the rate constant for spontaneous P-O cleavage of a phosphodiester anion, the reaction that is catalyzed by many phosphodiesterases (4).More recently, a rate constant of 6 ϫ 10 Ϫ7 s Ϫ1 has been reported for the decomposition of thymidylyl-3Ј-5Ј-thymidine (TpT) at 80°C in 1 M KOH (5). Extrapolation of the results obtained earlier for dimethyl phosphate hydrolysis in neutral solution (4), to 80°C, would indicate a rate Ϸ10 5 -fold slower. That discrepancy might indicate a major role for catalysis by hydroxide, but the hydrolysis of another dialkyl phosphodiester, bis-3-(4-carboxyphenyl)neopentyl phosphate (Np* 2 P), in which ␥-branching of the leaving alcohol prevents C-O cleavage (Fig. 1), also proceeds Ϸ10 5 -fold more slowly in 1 M KOH (6).In an effort to resolve that discre...

No abstract

Block copolymer self-assembly is normally conducted via post-polymerization processing at high dilution. In the case of block copolymer vesicles (or "polymersomes"), this approach normally leads to relatively broad size distributions, which is problematic for many potential applications. Herein we report the rational synthesis of low-polydispersity diblock copolymer vesicles in concentrated solution via polymerization-induced self-assembly using reversible addition-fragmentation chain transfer (RAFT) polymerization of benzyl methacrylate. Our strategy utilizes a binary mixture of a relatively long and a relatively short poly(methacrylic acid) stabilizer block, which become preferentially expressed at the outer and inner poly(benzyl methacrylate) membrane surface, respectively. Dynamic light scattering was utilized to construct phase diagrams to identify suitable conditions for the synthesis of relatively small, low-polydispersity vesicles. Small-angle X-ray scattering (SAXS) was used to verify that this binary mixture approach produced vesicles with significantly narrower size distributions compared to conventional vesicles prepared using a single (short) stabilizer block. Calculations performed using self-consistent mean field theory (SCMFT) account for the preferred self-assembled structures of the block copolymer binary mixtures and are in reasonable agreement with experiment. Finally, both SAXS and SCMFT indicate a significant degree of solvent plasticization for the membrane-forming poly(benzyl methacrylate) chains.

RAFT dispersion polymerization of benzyl methacrylate (BzMA) has been used previously (E. R. Jones, et al., Macromolecules, 2012, 45, 5091) to prepare poly(2-(dimethylamino)ethyl methacrylate)-poly-(benzyl methacrylate) (PDMA-PBzMA) diblock copolymer nanoparticles in ethanol via polymerizationinduced self-assembly (PISA). However, the rate of polymerization was relatively slow, with incomplete monomer conversions being obtained when targeting higher mean degrees of polymerization (DP) even after 24 h at 70°C. Herein we examine the effect of the addition of up to 20% w/w water co-solvent on the kinetics of BzMA polymerization for this PISA formulation. Significantly faster polymerizations were observed: for a target DP of 200, 90% BzMA conversion was achieved within just 6 h in the presence of 20% w/w water, compared to only 35% conversion in anhydrous ethanol under the same conditions. This rate enhancement enables much higher mean DPs to be obtained for the core-forming PBzMA and is attributed to greater partitioning of the BzMA monomer within the particles, which increases the local monomer concentration. However, the presence of water adversely affected the evolution of copolymer morphology from spheres to worms to vesicles when employing a relatively short PDMA chain transfer agent, with only kinetically-trapped spheres being obtained at higher levels of added water. Aqueous electrophoresis studies indicate that the PDMA stabilizer chains acquired partial cationic charge in the presence of water. This leads to more efficient inter-particle repulsion, thus preventing the sphere-sphere fusion events required for an evolution in morphology. In summary, the addition of water to such PISA formulations allows the more efficient synthesis of spherical nanoparticles, but should be used with caution if either diblock copolymer worms or vesicles are desired.This journal is

The link between the cognitive deficit associated with Alzheimer type dementia and the loss of cholinergic function in the disease provides a basis for examining muscarinic agonists as potential therapeutic agents. This paper describes the design and synthesis of novel azabicyclic methyl esters as ligands for the muscarinic receptor. Replacement of the methyl ester by a 3-methyl-1,2,4-oxadiazole ring produces potent metabolically more stable muscarinic agonists capable of penetrating the central nervous system. These compounds generally show improved affinity relative to the corresponding methyl esters. 3-Methyl-1,2,4-oxadiazole 7b has an affinity 4 times that of acetylcholine. Receptor affinity is discussed in relation to the size and geometry of the azabicyclic ring and the electronic properties of the heteroaromatic ring.