Visible-Light Mediated Tryptophan Modification in Oligopeptides Employing Acylsilanes

A method for the selective tryptophan modification and labelling of tryptophan-containing peptides is described. Photoirradiation of acylsilanes generates reactive siloxycarbenes which undergo H−N-insertion into the indole moiety of tryptophan to give stable silyl protected hemiaminals. This method is successfully applied to chemically modify various tryptophan containing oligopeptides. The method enables the selective introduction of alkynes to peptides that are eligible for further alkyne-azide click chemistry. In addition, the dansyl fluorophore can be conjugated to a peptide using this approach.     

The application of graphene as a sample support in transmission electron microscopy

Transmission electron microscopy has witnessed rampant development and surging point resolution over the past few years. The improved imaging performance of modern electron microscopes shifts the bottleneck for image contrast and resolution to sample preparation. Hence, it is increasingly being realized that the full potential of electron microscopy will only be realized with the optimization of current sample preparation techniques. Perhaps the most recognized issues are background signal and noise contributed by sample supports, sample charging and instability. Graphene provides supports of single atom thickness, extreme physical stability, periodic structure, and ballistic electrical conductivity. As an increasing number of applications adapting graphene to their benefit emerge, we discuss the unique capabilities afforded by the use of graphene as a sample support for electron microscopy.     

Intracellular Photophysics of an Osmium Complex bearing an Oligothiophene Extended Ligand

This contribution describes the excited-state properties of an Osmium-complex when taken up into human cells. The complex 1 [Os(bpy)₂(IP-4T)](PF₆)₂ with bpy=2,2′-bipyridine and IP-4T=2-{5′-[3′,4′-diethyl-(2,2′-bithien-5-yl)]-3,4-diethyl-2,2′-bithiophene}imidazo[4,5-f][1,10]phenanthroline) can be discussed as a candidate for photodynamic therapy in the biological red/NIR window. The complex is taken up by MCF7 cells and localizes rather homogeneously within in the cytoplasm. To detail the sub-ns photophysics of 1 , comparative transient absorption measurements were carried out in different solvents to derive a model of the photoinduced processes. Key to rationalize the excited-state relaxation is a long-lived ³ILCT state associated with the oligothiophene chain. This model was then tested with the complex internalized into MCF7 cells, since the intracellular environment has long been suspected to take big influence on the excited state properties. In our study of 1 in cells, we were able to show that, though the overall model remained the same, the excited-state dynamics are affected strongly by the intracellular environment. Our study represents the first in depth correlation towards ex-vivo and in vivo ultrafast spectroscopy for a possible photodrug.     

Novel strategy for the preparation of 3-perfluoroalkylated-2H-indazole derivatives

A simple and novel methodology for the synthesis of 3-perfluoroalkylated-2H-indazole derivatives has been elaborated. The perfluoroalkylation of readily available 2-nitrobenzaldimines bearing electron donating groups was performed using the Ruppert-Prakash reagent and its analogues to afford α-difluoromethylated, α-trifluoromethylated and α-pentafluoroethylated benzylamines. A final reductive cyclization mediated by SnCl₂·2H₂O led to 2H-indazoles containing perfluoroalkyl groups via the generation of a new NN bond in moderate to good yields.     

Total Synthesis and Structure Correction of the Cyclic Lipodepsipeptide Orfamide A

A total synthesis of the cyclic lipodepsipeptide natural product orfamide A was achieved. By developing a synthesis format using an aminoacid ester building block and SPPS protocol adaptation, a focused library of target compounds was obtained, in high yield and purity. Spectral and LC-HRMS data of all library members with the isolated natural product identified the ⁵Leu residue to be d - and the 3’-OH group to be R-configured. The structural correction of orfamide A by chemical synthesis and analysis was confirmed by biological activity comparison in Chlamydomonas reinhardtii, which indicated compound configuration to be important for bioactivity. Acute toxicity was also found against Trypanosoma brucei, the parasite causing African sleeping sickness.     

A complete picture of protein unfolding and refolding in surfactants

Interactions between proteins and surfactants are of relevance in many applications including food, washing powder formulations, and drug formulation. The anionic surfactant sodium dodecyl sulfate (SDS) is known to unfold globular proteins, while the non-ionic surfactant octaethyleneglycol monododecyl ether (C₁₂E₈) can be used to refold proteins from their SDS-denatured state. While unfolding have been studied in detail at the protein level, a complete picture of the interplay between protein and surfactant in these processes is lacking. This gap in our knowledge is addressed in the current work, using the β-sheet-rich globular protein β-lactoglobulin (bLG). We combined stopped-flow time-resolved SAXS, fluorescence, and circular dichroism, respectively, to provide an unprecedented in-depth picture of the different steps involved in both protein unfolding and refolding in the presence of SDS and C₁₂E₈. During unfolding, core–shell bLG-SDS complexes were formed within ∼10 ms. This involved an initial rapid process where protein and SDS formed aggregates, followed by two slower processes, where the complexes first disaggregated into single protein structures situated asymmetrically on the SDS micelles, followed by isotropic redistribution of the protein. Refolding kinetics (>100 s) were slower than unfolding (<30 s), and involved rearrangements within the mixing deadtime (∼5 ms) and transient accumulation of unfolded monomeric protein, differing in structure from the original bLG-SDS structure. Refolding of bLG involved two steps: extraction of most of the SDS from the complexes followed by protein refolding. These results reveal that surfactant-mediated unfolding and refolding of proteins are complex processes with rearrangements occurring on time scales from sub-milliseconds to minutes.

The time-resolved study reveals several transition states during SDS-induced unfolding of the protein, as well as under refolding of the protein by the nonionic surfactant C₁₂E₈.     

Observation of inhomogeneity in the lipid composition of individual nanoscale liposomes

Liposomes, or vesicles, have been studied extensively both as models of biological membranes and as drug delivery vehicles. Typically it is assumed that all liposomes within the same preparation are identical. Here by employing pairs of fluorescently labeled lipids we demonstrated an up to 10-fold variation in the relative lipid composition of individual liposomes with diameters between 50 nm and 15 μm. Since the physicochemical properties of liposomes are directly linked to their composition, a direct consequence of compositional inhomogeneities is a polydispersity in the properties of the individual liposomes in an ensemble.     

A Chatt-Type Catalyst with One Coordination Site for Dinitrogen Reduction to Ammonia

With [Mo(N₂)(P₂^MePP₂^Ph)] the first Chatt-type complex with one coordination site catalytically converting N₂ to ammonia is presented. Employing SmI₂ as reductant and H₂O as proton source 26 equivalents of ammonia are generated. Analogous Mo⁰-N₂ complexes supported by a combination of bi- and tridentate phosphine ligands are catalytically inactive under the same conditions. These findings are interpreted by analyzing structural and spectroscopic features of the employed systems, leading to the conclusion that the catalytic activity of the title complex is due to the strong activation of N₂ and the unique topology of the pentadentate tetrapodal (pentaPod) ligand P₂^MePP₂^Ph. The analogous hydrazido(2-) complex [Mo(NNH₂)(P₂^MePP₂^Ph)](BAr^F)₂ is generated by protonation with HBAr^F in ether and characterized by NMR and vibrational spectroscopy. Importantly, it is shown to be catalytically active as well. Along with the fact that the structure of the title complex precludes dimerization this demonstrates that the corresponding catalytic cycle follows a mononuclear pathway. The implications of a PCET mechanism on this reactive scheme are considered.