Enhancing the catalytic repertoire of nucleic acids: a systematic study of linker length and rigidity

The incorporation of potentially catalytic groups in DNA is of interest for the in vitro selection of novel deoxyribozymes, A series of 10 C5-modified analogues of 2'-deoxyuridine triphosphate have been synthesised that possess side chains of differing flexibility and bearing a primary amino or imidazole functionality, For each series of nucleotide analogues differing degrees of flexibility of the C5 side chain was achieved through the use of alkynyl, alkenyl and alkyl moieties, The imidazole function was conjugated to these CS-amino-modified nucleotides using either imidazole 4-acetic acid or imidazole 4-acrylic acid (urocanic acid), The substrate properties of the nucleotides (fully replacing dTTP) with Taq polymerase during PCR have been investigated in order to evaluate their potential applications for in vitro selection experiments, 5-(3-Aminopropynyl)dUTP and 5-(E-3-aminopropenyl)dUTP and their imidazole 4-acetic acid- and urocanic acid-modified conjugates were found to be substrates, In contrast, C5-amino-modified dUTPs with alkane or Z-alkene linkers and their corresponding conjugates were not substrates, The incorporation of these analogues during PCR has been confirmed by inhibition of restriction enzyme digestion using XbaI and by mass spectrometry of the PCR products.

Lightly branched comb polyesters: application in fast drying solvent-borne coating formulations

Novel oxazoline-based comb-polymers possessing linoleyl or oleic side chains have been synthesized and used to produce low viscosity coatings. Inclusion of the polymers in model paint formulations results in coatings that exhibit faster drying times than commercially available alkyd resin formulations. The comb polymers were produced from diol substituted oxazoline monomers that were synthesized through a scalable, solvent free protocol and purified by simple recrystallisation. Co-polymerisation of the oxazolines with adipic acid at 160 °C in the bulk resulted in the targeted polyester comb type polymers. The polymers were soluble in a range of organic solvents and compatible with commercial alkyd resins. Model paint formulations containing up to 40 wt% of the linoleyl-based comb polymers exhibited a dramatic reduction in viscosity (from 35 to 13 Poise at 25 °C) with increasing quantities of polymer added. Dynamic mechanical analysis (DMA) studies revealed that the drying rate of the model paint formulations containing the comb polymers was enhanced when compared with that of commercial alkyd resins.

Determination of orientations of aromatic groups in self-assembled peptide fibrils by polarised Raman spectroscopy

In this paper we describe a novel combination of Raman spectroscopy, isotope editing and X-ray scattering as a powerful approach to give detailed structural information on aromatic side chains in peptide fibrils. The orientation of the tyrosine residues in fibrils of the peptide YTIAALLSPYS with respect to the fibril axis has been determined from a combination of polarised Raman spectroscopy and X-ray diffraction measurements. The Raman intensity of selected tyrosine bands collected at different polarisation geometries is related to the values and orientation of the Raman tensor for those specific vibrations. Using published Raman tensor values we solved the relevant expressions for both of the two tyrosine residues present in this peptide. Ring deuteration in one of the two tyrosine side chains allowed for the calculation to be performed individually for both, by virtue of the isotopic shift that eliminates band overlapping. Sample disorder was taken into account by obtaining the distribution of orientations of the samples from X-ray diffraction experiments. The results provide previously unavailable details about the molecular conformation of this peptide, and demonstrate the value of this approach for the study of amyloid fibrils.

Influence of elastase on alanine-rich peptide hydrogels

The self-assembly of the alanine-rich amphiphilic peptides Lys(Ala)6Lys (KA6K) and Lys(Ala)6Glu (KA6E)with homotelechelic or heterotelechelic charged termini respectively has been investigated in aqueous solution. These peptides contain hexa-alanine sequences designed to serve as substrates for the enzyme elastase. Electrostatic repulsion of the lysine termini in KA6K prevents self-assembly, whereas in contrast KA6E is observed, through electron microscopy, to form tape-like fibrils, which based on X-ray scattering contain layers of thickness equal to the molecular length. The alanine residues enable efficient packing of the side-chains in a beta-sheet structure, as revealed by circular dichroism, FTIR and X-ray diffraction experiments. In buffer, KA6E is able to form hydrogels at sufficiently high concentration. These were used as substrates for elastase, and enzyme-induced de-gelation was observed due to the disruption of the beta-sheet fibrillar network. We propose that hydrogels of the simple designed amphiphilic peptide KA6E may serve as model substrates for elastase and this could ultimately lead to applications in biomedicine and regenerative medicine.

Enzymatically-triggered, isothermally responsive polymers: re-programming poly(oligoethylene glycols) to respond to phosphatase

Polymers which can respond to externally applied stimuli have found much application in the biomedical field due to their (reversible) coil–globule transitions. Polymers displaying a lower critical solution temperature are the most commonly used, but for blood-borne (i.e., soluble) biomedical applications the application of heat is not always possible, nor practical. Here we report the design and synthesis of poly(oligoethylene glycol methacrylate)-based polymers whose cloud points are easily varied by alkaline phosphatase-mediated dephosphorylation. By fine-tuning the density of phosphate groups on the backbone, it was possible to induce an isothermal transition: A change in solubility triggered by removal of a small number of phosphate esters from the side chains activating the LCST-type response. As there was no temperature change involved, this serves as a model of a cell-instructed polymer response. Finally, it was found that both polymers were non cytotoxic against MCF-7 cells (at 1 mg·mL–1), which confirms promise for biomedical applications.

Nanozipper formation in the solid state from a self-assembling tripeptide with a single tryptophan residue

The self-assembly of a terminally protected tripeptide Boc-gamma-Abu(1)-Ala(2)-Trp(3)-OMe (gamma-Abu = gamma-aminobutyric acid) I results in the formation of a nanostructured supramolecular zipper through various non-covalent interactions in the crystal in which the indole side-chain of the Trp(3) residue plays a key role via N-H...pi interactions. (c) 2006 Published by Elsevier Ltd.