Mechanochemical Formation of Polyacrylamide Nanocomposites: Impact of Nanoparticle Surface Chemistry and Substrate Functionalization

Osteoarthritis is a degenerative joint condition, with treatments currently limited to symptom management and invasive surgical procedures. This project investigates the fundamental mechanisms for a potential treatment route that would allow soft material nanocomposites to form within the joint through mechanochemical reactions. To gain fundamental insight into the different chemical pathways for this to occur, this work compares a top-down vs bottom-up approach. The top-down method focuses on the influence of applied oscillatory mechanical stresses on bulk hydrogel polymerization. Polyacrylamide (PAM) is used as a well-studied control system. PAM-nanocomposites were achieved through co-polymerization, with gold nanoparticles with different capping ligands directly added to the PAM precursor solution. A rheometer was used to simultaneously apply the small angle oscillatory shear and monitor gelation time. Initial results indicate decreased gelation time for PAM-nanocomposites with gold nanoparticle capping ligands of citrate and CTAB (cetyltrimethylammonium), relative to no nanoparticles present. Accelerated gelation times, relative to no nanoparticles present, were observed for PVP (polyvinylpyrrolidone) and PAA (polyacrylic acid) capping ligands. Comparing molecular structure vs chemical functionalities, these results suggest structure (e.g. polymer-based capping ligands, PVP and PAA) over functionality (e.g. hydrogen bonding for PAA and citric acid, but not CTAB and to a lesser degree PVP) influences gelation time. The complementary bottom-up method uses a 3-metharcyloxypropyltrimethoxysilane functionalized silica surface and AFM as a nanoscale single point sliding contact to grow a surface bound PAM hydrogel film. Future work will continue to quantify these pathways, examining reactions within a stress-assisted Arrhenius model

Role of Intermolecular Interactions between Nanoparticle Capping Ligands and Hydrogel Surfaces During Sliding

Modern treatments for osteoarthritis increasingly involve the use of nanoparticles as drugdelivery systems, but there is little known about the influence of nanoparticle chemical composition and surface chemistry between nanoparticles and soft materials in sliding contact. This work implements cartilage-mimicking polyacrylamide (PAM) hydrogels as a well-studied, fundamental platform. In situ (in a fluid environment) macroscale friction tests as a function of shear rate were conducted with a rheometer with a tribology adapter, controlling for contact pressure. Comparing different nanoparticle compositions, citrate capped metal (gold) nanoparticles exhibited a 50% increase in friction relative to water. With no difference in solution viscosity, this difference is likely driven by hydrogen bonding between the citrate ligands and PAM surface. In contrast, carbon based nanoparticles (nanodiamonds) with no capping ligands exhibited a 50% decrease in friction relative to water. Here, a higher solution viscosity for the nanodiamonds is likely dictating the sliding mechanism. Additional tests exploring gold nanoparticles with controlled capping ligands further support the impact of intermolecular interactions between nanoparticle capping ligands and the PAM surface in controlling sliding mechanisms. Post-sliding characterization of the PAM with confocal Raman microscopy surfaces indicate no damage to the hydrogel, and the presence of uncapped nanoparticle aggregates. Next steps will focus on the extent to which nanoparticles might be embedded within the PAM surface as a result of sliding

Crystallization and preliminary X-ray analysis of the 9 kDa protein of the mouse signal recognition particle and the selenomethionyl-SRP9

AbstractTwo different crystal forms of the 9 kDa protein of the signal recognition particle (SRP9) have been prepared by the hanging drop vapor diffusion technique using 28% (w/v) PEG8000 or 28% saturated ammonium sulphate as precipitant. The crystals are hexagonal bipyramids with average dimensions of 0.2 × 0.1 × 0.1 mm3 and they diffract to a resolution of 2.3 Å. They belong to the space groups P6222/P6422 or P3121/P3221 with cell dimensions a = b = 63.0 Å, and c = 111.5 Å. Crystals have also been grown from the selenomethionyl protein and multiwavelength data sets have been collected

Alu elements as regulators of gene expression

Alu elements are the most abundant repetitive elements in the human genome; they emerged 65 million years ago from a 5′ to 3′ fusion of the 7SL RNA gene and amplified throughout the human genome by retrotransposition to reach the present number of more than one million copies. Over the last years, several lines of evidence demonstrated that these elements modulate gene expression at the post-transcriptional level in at least three independent manners. They have been shown to be involved in alternative splicing, RNA editing and translation regulation. These findings highlight how the genome adapted to these repetitive elements by assigning them important functions in regulation of gene expression. Alu elements should therefore be considered as a large reservoir of potential regulatory functions that have been actively participating in primate evolution

Tool Workshop

This all-day workshop provides a setting in which participants learn to use handheld instruments, conduct a building performance case study through structured methodology, develop hypothesis and investigation strategy formulation, are allowed behind-the-scenes building investigations, and make team presentations of study results to the workshop participants. Participants gain understanding of objective and subjective procedures for performing post-occupancy evaluations of building performance

The signal recognition particle and related small cytoplasmic ribonucleoprotein particles

Recently, a number of novel small cytoplasmic ribonucleoprotein particles have been identified that comprise RNA and protein subunits related to the signal recognition particle (SRP). Here we discuss the latest results on the structure and functions of SRP together with the structures and putative functions of the novel SRP-related ribonucleoprotein particles

The heterodimeric subunit SRP9/14 of the signal recognition particle functions as permuted single polypeptide chain.

The targeting of nascent polypeptide chains to the endoplasmic reticulum is mediated by a cytoplasmic ribonucleoprotein, the signal recognition particle (SRP). The 9 kD (SRP9) and the 14 kD (SRP14) subunits of SRP are required to confer elongation arrest activity to the particle. SRP9 and SRP14 form a heterodimer which specifically binds to SRP RNA. We have constructed cDNAs that encode single polypeptide chains comprising SRP9 and SRP14 sequences in the two possible permutations linked by a 17 amino acid peptide. We found that both fusion proteins specifically bound to SRP RNA as monomeric molecules folded into a heterodimer-like structure. Our results corroborate the previous hypothesis that the authentic heterodimer binds to SRP RNA in equimolar ratio. In addition, both fusion proteins conferred elongation arrest activity to SRP(-9/14), which lacks this function, and one fusion protein could functionally replace the heterodimer in the translocation assay. Thus, the normal N-and C-termini of both proteins have no essential role in folding, RNA-binding and in mediating the biological activities. The possibility to express the heterodimeric complex as a single polypeptide chain facilitates the analysis of its functions and its structure in vivo and in vitro

The SRP9/14 subunit of the human signal recognition particle binds to a variety of Alu-like RNAs and with higher affinity than its mouse homolog.

The heterodimeric subunit, SRP9/14, of the signal recognition particle (SRP) has previously been found to bind to scAlu and scB1 RNAs in vitro and to exist in large excess over SRP in anthropoid cells. Here we show that human and mouse SRP9/14 bind with high affinities to other Alu-like RNAs of different evolutionary ages including the neuron-specific BC200 RNA. The relative dissociation constants of the different RNA-protein complexes are inversely proportional to the evolutionary distance between the Alu RNA species and 7SL RNA. In addition, the human SRP9/14 binds with higher affinity than mouse SRP9/14 to all RNAs analyzed and this difference is not explained by the additional C-terminal domain present in the anthropoid SRP14. The conservation of high affinity interactions between SRP9/14 and Alu-like RNAs strongly indicates that these Alu-like RNPs exist in vivo and that they have cellular functions. The observation that human SRP9/14 binds better than its mouse counterpart to distantly related Alu RNAs, such as recently transposed elements, suggests that the anthropoid-specific excess of SRP9/14 may have a role in controlling Alu amplification rather than in compensating a defect in SRP assembly and functions