ChemBioChem

The cover picture shows a photoswitchable affinity label (PAL) in its trans form bound noncovalently to bovine carbonic anhydrase 2. Potential reactive nucleophiles surrounding the PAL are highlighted in yellow. Histidines 2 and 3, which eventually undergo the bioconjugation, are shown in red. After bioconjugation, the affinity label can be withdrawn from the active site through photoisomerization (upper left corner). For details see the article by J. H. Harvey and D. Trauner on p. 191 ff.

Fathoming fibrils. Solid-state NMR spectroscopy is becoming a powerful tool for the structure elucidation of amyloid fibrils. An overview of solid-state NMR spectroscopic techniques for structure elucidation in proteins, and a review of recent results that have been obtained by solid-state NMR spectroscopy and other techniques on selected amyloid systems is presented here. Finally, an overview of the structural motifs that have been identified as amyloid-forming elements is presented.

Best PALs. We have applied a photoswitchable affinity label (PAL) to the optical control of protein function. The enzymatic activity of native carbonic anhydrase was controlled with covalently tethered, photoswitchable inhibitors (see scheme). This system allows the photoregulation of native proteins without site-specific introduction of highly reactive residues.

Let's go round. We report the development of a transposition based genetic selection methodology used to uncover three cyclic peptide inhibitors of the E. coli methyltransferase. The activity of the selected cyclic peptides was confirmed in vivo and in vitro. The IC₅₀ of the most active cyclic peptide (SGWYVRNM, shown in the figure) was comparable to that of the known methyltransferase inhibitor, sinefungin.

A good GAG. In order to modulate glycosaminoglycan (GAG) biosynthesis, and determine their physiological and developmental role in a spatiotemporal manner, we synthesized a library of “click” xylosides with various aglycone moieties that are very stable under physiological conditions. The synthesized compounds were examined for their ability to stimulate GAG chains in a cell line that is defective in the production of GAG chains.

Plastics provide a moldable platform for molecular analyses. A system was developed that permitted receptors to be doped within moldable polycarbonate by using a dual anchor–receptor design. These resins were molded into 3D objects without loss in the affinity of a pendant receptor. This finding demonstrates a practical advance for the display of molecular species that allows one to mold affinity-based systems into objects from the micron to meter scale.

Flick of the switch. We used an aptazyme and an anti-ribosome binding site (RBS) to construct a novel artificial ribozyme-based riboswitch that functions in E. coli (see figure). This riboswitch worked well at low temperature and activated its own ribozyme and gene expression in the presence of targets even though it is ribozyme based. We also improved the switching efficiency by constructing a cascading system.

Flip the switch. Riboswitches, which are RNA sequences that undergo a ligand-induced conformational change to alter gene expression, can be used to reprogram how bacteria respond to small molecules. Here we present a new selection method based on cell motility (outlined in the figure) that rapidly identifies synthetic riboswitches and minimizes cost and effort.

Don't be so negative. We have synthesized new dichlorofluorescein derivatives that are less negatively charged than fluorescein. The synthesis is concise and scalable, and the compounds are conjugation-ready. In addition, these compounds permeate zebrafish embryos; this suggests that they might be useful fluorescent probes for bioimaging.

Target: central nervous system. Cationic liposome–plasmid DNA complexes (see figure) were modified with a PEG stealth layer, before the addition of C-terminal fragments of tetanus toxin (TH_C), botulinum toxin (BH_C) or the truncated C-terminal domain of TH_C (TH_CC) as biological “targeting” ligands. Transfection of neuronal cell lines with these vectors were up to 30-fold higher than control transfections with nanoparticles that lacked the protein ligand.

Two in one. A bifunctional nucleic acid that consists of the cocaine and adenosine 5′-monophosphate (AMP) aptamers is blocked by a nucleic acid. The complex is separated by either cocaine or AMP, and the process is sensed by a DNAzyme colorimetric reaction or by the electronic transduction by using impedance spectroscopy or a field-effect transistor.

The solution conformation of a dermatan-derived tetrasaccharide—ΔHexA-(1→3)-GalNAc4S-β-(1→4)-IdoA-α-(1→3)-red-GalNAc4S—has been explored by means of NMR spectroscopy, including NOE and RDC analysis, assisted by MD simulations. The investigation confirmed a moderate flexibility of the two external anomeric torsions, while highlighting a certain degree of flexibility of the central glycosidic linkage.

The magnificent seven. New polyhydroxylated azepanes have been designed as stable mimics of 1-azasugar glycosidase inhibitors that include noeuromycin. Some of these compounds display potent and selective competitive inhibition towards glycosidases, as well as some activity towards glucosylceramide transferase involved in Gaucher's disease.

Handle with care. The ability of a mutant P450_BM3 that lacks a catalytically important threonine residue (Thr268Ala) to hydroxylate fatty acids has been found to be dependant upon the chain length of the substrate (see graph). This indicates that caution is required in assuming a loss of activity with such threonine to alanine mutants of P450 enzymes, as activity with certain substrates might be maintained.

Mannopyranoside oligomers. A series of synthetic α-(1→6)-linked octyl mannopyranoside oligomers and methoxy and deoxy disaccharide analogues were evaluated to explore the acceptor specificity of a polyprenol monophosphomannose-dependent α-(1→6)-mannosyltransferase that is involved in the biosynthesis of the mannan core of mycobacterial lipoarabinomannan.

Going out in style. The quenching of the fluorescent nucleobases benzopyrene, perylene, and pyrene by neighboring natural bases is described, as well as a strategy for insulating fluorophores from PET quenching.

Membrane depolarization. Lipophilic molecular probes with 2,6-bis(zinc(II)-dipicolylamine)phenoxide as the molecular recognition unit induce membrane transport and are toxic to mammalian cells. A hydrophilic analogue is selectively active against drug-resistant strains of Staphylococcus aureus.

It's all about presentation. Neoglycoprotein antigens that bear synthetic, complex oligosaccharides target dendritic cell-surface receptors and cause a 50-fold enhancement of antigen uptake and presentation to CD4⁺ T cells. A tenfold enhancement is observed for CD8⁺ T cells; this indicates that the targeted lectin(s) can mediate cross-presentation of antigens on MHC class I. The observed enhancements are unique to complex oligosaccharides.

Into the fold. The cancer biomarker HMGA1a protein was found to function like a chaperone for synthetic foldable polymers, and was able to manipulate the nanostructures of the polymers by unfolding and refolding them. This protein switched on either blue or yellow fluorescence depending solution properties; conversely, the polymer behaved as a unique two-way bioindicator for the cancer biomarker HMGA1a.

Possible in vivo redox-controlled pathway for denitration of nitrated proteins. Hemoglobin not only catalyzes H₂O₂/nitrite-dependent self-nitration and oxidation, but also mediates nitro reduction of 3-nitrotyrosine to 3-aminotyrosine in human hemoglobin in the presence of antioxidants in a dose-dependent fashion. Mass spectrometry provides convincing evidence for H₂O₂/nitrite-mediated post-translational modifications of hemoglobin and redox-regulated tyrosine nitration.

On or off. The processing of N-terminal unnatural amino acids (UAAs) in E. coli was studied. Controlled retention or cleavage of the N-terminal UAA in a nascent polypeptide was achieved by selecting an appropriate penultimate amino acid (represented by dark blue rectangles), as illustrated in the scheme.