Fr Francium

The inside cover picture shows a rationally designed MD2 hot-spot-mimicking peptide in complex with toll-like receptor 4, which is an important cell-surface receptor that plays critical roles in the innate immune system and in inflammation response. For further details on the binding affinity and specificity of this truncated peptide, see the paper by H. Yin et al. on p. 1827 ff.

The cover picture shows a model of a luminescent iridium(III) complex functionalized with three Ac-TZ14011 peptide moieties binding to the chemokine receptor 4 (CXCR4). The G protein-coupled receptor CXCR4 is over-expressed in many types of cancer and, therefore, is a potential biomarker in the field of diagnostic oncology. In the CPK model, the three Ac-TZ14011 peptides are colored by element and the iridium complex is red. The iridium complex has a luminescence lifetime in the order of 200 ns; this enables visualization with fluorescence lifetime imaging microscopy (FLIM). The FLIM image of cells incubated with the peptide–iridium conjugate in the background shows accumulation in the cell membranes; this is in accordance with the membrane location of CXCR4. For further information, see the communication by A. H. Velders, F. W. B. van Leeuwen, et al. on p. 1897 ff.

Peptide-bond isosteres can enable a deep interrogation of the structure and function of a peptide or protein by amplifying or attenuating particular chemical properties. In this Minireview, the electronic, structural, and conformational attributes of four such isosteres—thioamides, esters, alkenes, and fluoroalkenes—are examined in detail. In particular, the ability of these isosteres to partake in noncovalent interactions is compared with that of the peptide bond. The consequential perturbations provide a useful tool for chemical biologists to reveal new structure–function relationships, and to endow peptides and proteins with desirable attributes.

Not better, just different: The electronic, structural, and conformational attributes of four peptide-bond isosteres—thioamides, esters, alkenes, and fluoroalkenes—can enable chemical biologists to probe structure–function relationships of peptides and proteins with unprecedented precision.

Hsp70 is biologically relevant for its chaperon functions. The CX⁻ and CX⁺ sublines, which derive from the parental colon carcinoma CX2 cell line, are accordingly very similar. They have been reported to be specifically different only in Hsp70 membrane expression, which is associated with immunostimulatory effects. CX⁻/CX⁺ have been phenotypically characterized by immunofluorescence studies and Raman spectroscopy combined with robust clustering and multivariate analysis. With the latter we address the potential of overall characterization for CX⁻/CX⁺ discrimination and gain molecular insights into Hsp70 differential expression. Due to their strong resemblance, CX⁻ and CX⁺ show similar mean Raman spectra, which look indiscernible at first. Interestingly, their rather protein-dominated Raman spectra reveal, besides changes in protein and amino acids, very specific changes in DNA/RNA nucleotides involving pyrimidine ring Raman hypochromic effects. Therefore, discriminating CX⁻ from CX⁺ is ultimately achieved based on principal component scores. Because CX⁻/CX⁺ are associated with the same lipid marker, changes in proteins support lipid interactions with regulatory proteins. More importantly, changes observed in nucleobases, which are indicative of DNA/RNA–protein binding interactions, suggest transcription deregulations as participating precursor onsets of different transport mechanisms that lead to Hsp70 differential expression and associated phenotypic variation. Besides immunofluorescence, we have used Raman spectroscopy combined with multivariate analysis within an autologous tumor system for label-free nondestructive cell-subline discrimination, and demonstrate, to our knowledge, the first overall phenotypic monitoring with insights into Hsp70 differential expression. This might well prove to be useful for Raman label-free cell-sorting of the CX⁻/CX⁺ sublines.

Split screen: The picture shows colon carcinoma CX2 line expansion into the sublines CX⁻/CX⁺ due to their specifically different Hsp70 expression patterns. Only immunofluorescence helps to quantify Hsp70 in order to FACS-sort the cells. Yet, Raman fingerprints of CX⁻/CX⁺ not only hold potential for label-free nondestructive cell sorting, but could also provide insights into nucleobase changes attributed to alterations in DNA binding interactions.

Native chemical ligation:N-sulfanylethylanilide (SEAlide) peptides act as thioester peptides in the presence of N-terminal cysteinyl peptides and an appropriate choice of salts. With the use of the SEAlide peptide, a kinetically controlled native chemical ligation proceeding in an unprecedented highly selective manner was achieved. Additionally, a one-pot, four-component coupling under kinetic conditions was achieved.

Tuberactinomycin biosynthesis: Viomycin and capreomycin are structurally related antituberculosis drugs. Analysis of the nonribosomal peptide synthetases that assemble these molecules determined that peptide assembly occurs by distinct mechanisms. These unexpected differences provide insights into how modifications to homologous nonribosomal peptide synthetases can result in structure diversification.

The chemokine receptor 4 (CXCR4) is over-expressed in 23 types of cancer in which it plays a role in, among others, the metastatic spread. For this reason it is a potential biomarker for the field of diagnostic oncology. The antagonistic Ac-TZ14011 peptide, which binds to CXCR4, has been conjugated to luminescent iridium dyes to allow for CXCR4 visualization. The iridium dyes are cyclometalated octahedral iridium(III) 2-phenylpyridine complexes that can be functionalized with one, two or three targeting Ac-TZ14011 peptides. Confocal microscopy and fluorescence lifetime imaging microscopy (FLIM) showed that the peptide–iridium complex conjugates can be used to visualize CXCR4 expression in tumor cells. The CXCR4 receptor affinity and specific cell binding of the mono-, di- and trimeric peptide derivatives were assessed by using flow cytometry. The three derivatives possessed nanomolar receptor affinity and could distinguish between cell lines with different CXCR4 expression levels. This yields the first example of a neutral iridium(III) complex functionalized with peptides for FLIM-based visualization of a cancer associated membrane receptor.

FLIM festival: The chemokine receptor 4 (CXCR4) is over-expressed in 23 types of cancer. Different numbers of an antagonistic CXCR4-targeting peptide have been conjugated to luminescent iridium complexes. We have found that these conjugates are able bind CXCR4 with high affinity (see figure), and can be used in fluorescence lifetime imaging microscopy (FLIM) experiments.