ChemBioChem

Cationic Polymers with Inhibition Ability of DNA Condensation Elevate Gene Expression.

Publication Date: 2010 Aug 30 PMID: 20806309
Authors: Harada, A. - Kimura, Y. - Kono, K.
Journal: Chembiochem

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Spontaneous Protein Crowding in Liposomes: A New Vista for the Origin of Cellular Metabolism.

Publication Date: 2010 Aug 30 PMID: 20806308
Authors: Luisi, P. L. - Allegretti, M. - Pereira de Souza, T. - Steiniger, F. - Fahr, A. - Stano, P.
Journal: Chembiochem

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Multifaceted Modes of Action for the Glutarimide-Containing Polyketides Revealed.

Publication Date: 2010 Aug 30 PMID: 20806307
Authors: Rajski, S. R. - Shen, B.
Journal: Chembiochem

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Divergent Heparin-Induced Fibrillation Pathways of a Prion Amyloidogenic Determinant.

Publication Date: 2010 Aug 26 PMID: 20799315
Authors: Bazar, E. - Jelinek, R.
Journal: Chembiochem

Polysaccharides and glycosaminoglycans (GAGs), particularly heparin, have been shown to directly affect fibrillation phenomena and the biological activities of amyloid proteins. We present a systematic analysis of the impact of heparin upon fibrillation of the amyloidogenic determinant of the prion protein PrP(106-126). Experimental data, including thioflavin T fluorescence, transmission electron microscopy, and circular dichroism, demonstrate that heparin induced dramatically diverging aggregation pathways of PrP(106-126). Specifically, enhanced beta-sheet formation of the prion fragment leading to fibril assemblies occurred in solutions containing low heparin/prion mole ratios, while mixtures containing a greater abundance of heparin showed almost complete inhibition of PrP(106-126) fibril formation. Based upon the experimental data we have proposed a unified model accounting for the interplay between the roles of heparin as a scaffold for nucleation and fibril growth on the one hand and as a disruptor of fibrillation through electrostatic affinity with the monomeric peptide units on the other. This study clarifies previous conflicting studies, and concludes that GAGs inhibit fibrillation and amyloid toxicity in some cases, and promote amyloidogenesis in others.

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A DNA Polymerase with Increased Reactivity for Ribonucleotides and C5-Modified Deoxyribonucleotides.

Publication Date: 2010 Aug 23 PMID: 20734370
Authors: Staiger, N. - Marx, A.
Journal: Chembiochem

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Probing the Micelle-Bound Aggregation-Prone State of alpha-Synuclein with (19)F NMR Spectroscopy.

Publication Date: 2010 Aug 20 PMID: 20730847
Authors: Wang, G. F. - Li, C. - Pielak, G. J.
Journal: Chembiochem

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Catalysis by Dihydrofolate Reductase from the Psychropiezophile Moritella profunda.

Publication Date: 2010 Aug 19 PMID: 20726028
Authors: Evans, R. M. - Behiry, E. M. - Tey, L. H. - Guo, J. - Loveridge, E. J. - Allemann, R. K.
Journal: Chembiochem

The influence of temperature and pH on the stability and catalytic activity of dihydrofolate reductase (MpDHFR) from the cold-adapted deep-sea bacterium Moritella profunda was studied. The thermal melting temperature was found to be ~38 degrees C and was not affected by pH, while activity measurements demonstrated that its stability was maximal at pH 7 and was reduced dramatically below pH 6 or above pH 8. The steady-state rate constant (k(cat)) was maximal at neutral pH and higher temperatures, while the Michaelis constants (K(M)) for both substrate and cofactor were optimal at lower temperatures and at elevated or reduced pH. For both temperature and pH, any change in k(cat) was therefore offset by a similar change in K(M). Both the activation enthalpy and entropy of the MpDHFR-catalysed reaction were lower than those of DHFR from E. coli leading overall to a very small difference in activation free energy and therefore similar steady-state rate constants at the same temperature. The chemical step of the reaction is not rate limiting at pH 7, but becomes progressively more rate limiting as the pH increases. These results demonstrate adaptation of MpDHFR to its environment and show compromises between enthalpic and entropic contributions to the reaction, and between k(cat) and K(M).

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Preparation of a Functional GABARAP-Lipid Conjugate in Nanodiscs and its Investigation by Solution NMR Spectroscopy.

Publication Date: 2010 Aug 16 PMID: 20715272
Authors: Ma, P. - Mohrluder, J. - Schwarten, M. - Stoldt, M. - Singh, S. K. - Hartmann, R. - Pacheco, V. - Willbold, D.
Journal: Chembiochem

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Membrane-Surface Anchoring of Charged Diacylglycerol-Lactones Correlates with Biological Activities.

Publication Date: 2010 Aug 16 PMID: 20715268
Authors: Raifman, O. - Kolusheva, S. - El Kazzouli, S. - Sigano, D. M. - Kedei, N. - Lewin, N. E. - Lopez-Nicolas, R. - Ortiz-Espin, A. - Gomez-Fernandez, J. C. - Blumberg, P. M. - Marquez, V. E. - Corbalan-Garcia, S. - Jelinek, R.
Journal: Chembiochem

Synthetic diacylglycerol-lactones (DAG-lactones) are effective modulators of critical cellular signaling pathways, downstream of the lipophilic second messenger diacylglycerol, that activate a host of protein kinase C (PKC) isozymes and other nonkinase proteins that share similar C1 membrane-targeting domains with PKC. A fundamental determinant of the biological activity of these amphiphilic molecules is the nature of their interactions with cellular membranes. This study examines the biological properties of charged DAG-lactones exhibiting different alkyl groups attached to the heterocyclic nitrogen of an alpha-pyridylalkylidene chain, and particularly the relationship between membrane interactions of the substituted DAG-lactones and their respective biological activities. Our results suggest that bilayer interface localization of the N-alkyl chain in the R(2) position of the DAG-lactones inhibits translocation of PKC isoenzymes onto the cellular membrane. However, the orientation of a branched alkyl chain at the bilayer surface facilitates PKC binding and translocation. This investigation emphasizes that bilayer localization of the aromatic side residues of positively charged DAG-lactone derivatives play a central role in determining biological activity, and that this factor contributes to the diversity of biological actions of these synthetic biomimetic ligands.

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Analysis of the Sorangicin Gene Cluster Reinforces the Utility of a Combined Phylogenetic/Retrobiosynthetic Analysis for Deciphering Natural Product Assembly by trans-AT PKS.

Publication Date: 2010 Aug 16 PMID: 20715267
Authors: Irschik, H. - Kopp, M. - Weissman, K. J. - Buntin, K. - Piel, J. - Muller, R.
Journal: Chembiochem

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Solution Structure of the Leader Sequence of the Patellamide Precursor Peptide, PatE(1-34).

Publication Date: 2010 Aug 16 PMID: 20715266
Authors: Houssen, W. E. - Wright, S. H. - Kalverda, A. P. - Thompson, G. S. - Kelly, S. M. - Jaspars, M.
Journal: Chembiochem

The solution structure of the leader sequence of the patellamide precursor peptide was analysed by using CD and determined with NOE-restrained molecular dynamics calculations. This leader sequence is highly conserved in the precursor peptides of some other cyanobactins harbouring heterocycles, and is assumed to play a role in targeting the precursor peptide to the post-translational machinery. The sequence was observed to form an alpha-helix spanning residues 13-28 with a hydrophobic surface on one side of the helix. This hydrophobic surface is proposed to be the site of the initial binding with modifying enzymes.

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Novel Hybrid Esterase-Haloacid Dehalogenase Enzyme.

Publication Date: 2010 Aug 16 PMID: 20715265
Authors: Beloqui, A. - Polaina, J. - Vieites, J. M. - Reyes-Duarte, D. - Torres, R. - Golyshina, O. V. - Chernikova, T. N. - Waliczek, A. - Aharoni, A. - Yakimov, M. M. - Timmis, K. N. - Golyshin, P. N. - Ferrer, M.
Journal: Chembiochem

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Modulation of Shank3 PDZ Domain Ligand-Binding Affinity by Dimerization.

Publication Date: 2010 Aug 16 PMID: 20715264
Authors: Iskenderian-Epps, W. S. - Imperiali, B.
Journal: Chembiochem

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Fluorous Iminoalditols: A New Family of Glycosidase Inhibitors and Pharmacological Chaperones.

Publication Date: 2010 Aug 16 PMID: 20715263
Authors: Schitter, G. - Steiner, A. J. - Pototschnig, G. - Scheucher, E. - Thonhofer, M. - Tarling, C. A. - Withers, S. G. - Fantur, K. - Paschke, E. - Mahuran, D. J. - Rigat, B. A. - Tropak, M. B. - Illaszewicz, C. - Saf, R. - Stutz, A. E. - Wrodnigg, T. M.
Journal: Chembiochem

A collection of new reversible glycosidase inhibitors of the iminoalditol type featuring N-substituents containing perfluorinated regions has been prepared for evaluation of physicochemical, biochemical and diagnostic properties. The vast variety of feasible oligofluoro moieties allows for modular approaches to customised structures according to the intended applications, which are influenced by the fluorine content as well as the distance of the fluorous moiety from the ring nitrogen. The first examples, in particular in the D-galacto series, exhibited excellent inhibitory activities. A preliminary screen with two human cell lines showed that, at subinhibitory concentrations, they are powerful pharmacological chaperones enhancing the activities of the catalytically handicapped lysosomal D-galactosidase mutants associated with GM1 gangliosidosis and Morquio B disease.

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Precision Control of Cellular Pathways with Light.

Publication Date: 2010 Aug 4 PMID: 20687052
Authors: Lemke, E. A.
Journal: Chembiochem

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Influence of cleavage site on global folding of an RNA-cleaving DNAzyme.

Publication Date: 2010 Aug 16 PMID: 20665772
Authors: Lam, J. C. - Li, Y.
Journal: Chembiochem

8-17 is a DNAzyme with metal-dependent endoribonuclease activity. Recently, a variant termed 8-17NG was reported as the first nucleic acid enzyme capable of cleaving all 16 dinucleotide junctions of RNA with rate enhancements ranging from 1000- to 1,000,000,000-fold over background activity. We attributed this broad-ranging cleavage efficiency to global folding of the DNAzyme. We sought to examine the influence of dinucleotides at the cleavage site of 8-17NG on global folding by using three-color (3c) FRET. By comparing the folding of 8-17NG with all 16 possible dinucleotide junctions, we found all examined DNAzyme-substrate constructs adopted a two-step folding process in the presence of Mn(2+), which was consistent with previous metal-induced folding studies of 8-17. Interestingly, Mn(2+) titration experiments also suggest that the second folding step is dependent on dinucleotide identity: purine-purine junctions allowed 8-17NG to fold at lower concentrations than pyrimidine-pyrimidine linkages. This finding was corroborated by RNA cleavage assays, in which the largest improvement in cleavage yield was observed in pyrimidine-pyrimidine junctions when [Mn(2+)] was increased. Taken together, these results support the previously observed hierarchy of 8-17 activity for different cleavage sites. Complemented by earlier sequence and structure-function studies, this investigation allowed for the first detailed examination of crucial relationships between the structural influence and junction preferences of nucleic acid-catalyzed RNA cleavage reactions.

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Cellular internalization of water-soluble helical aromatic amide foldamers.

Publication Date: 2010 Aug 16 PMID: 20665771
Authors: Iriondo-Alberdi, J. - Laxmi-Reddy, K. - Bouguerne, B. - Staedel, C. - Huc, I.
Journal: Chembiochem

The intracellular transport of drugs and therapeutics represents one of the most exciting and challenging areas at the interface of chemistry, biology, and medicine. Most of the effort in this field so far has been devoted to the development of peptide-based delivery systems that can translocate therapeutic agents into their intracellular targets. More recently, the use of bioinspired non-natural foldamers has resulted in the successful delivery of cargo molecules, which possess a wide range of sizes and physicochemical properties across the cell membrane. We report herein the synthesis of aromatic amide foldamers and their biological evaluation as cell-penetrating agents. By using a well-established synthetic route, a series of fluorescein-labeled cationic aryl amide conjugates has been constructed, and their cellular uptake into various human cell lines has been analyzed by flow cytometry and fluorescence microscopy. The assays revealed that longer oligomers achieve greater cellular translocation, with octamer Q8 proving to be a remarkable vehicle for all three cell lines. Biological studies have also indicated that these helices are biocompatible, thus showing promise in their application as cell-penetrating agents and as vehicles to deliver biologically active molecules into cells.

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Pacidamycin biosynthesis: identification and heterologous expression of the first uridyl peptide antibiotic gene cluster.

Publication Date: 2010 Aug 16 PMID: 20665770
Authors: Rackham, E. J. - Gruschow, S. - Ragab, A. E. - Dickens, S. - Goss, R. J.
Journal: Chembiochem

The pacidamycins are antimicrobial nucleoside antibiotics produced by Streptomyces coeruleorubidus that inhibit translocase I, an essential bacterial enzyme yet to be clinically targeted. The novel pacidamycin scaffold is composed of a pseudopeptide backbone linked by a unique exocyclic enamide to an atypical 3'-deoxyuridine nucleoside. In addition, the peptidyl chain undergoes a double inversion caused by the incorporation of a diamino acid residue and a rare internal ureido moiety. The pacidamycin gene cluster was identified and sequenced, thereby providing the first example of a biosynthetic cluster for a member of the uridyl peptide family of antibiotics. Analysis of the 22 ORFs provided an insight into the biosynthesis of the unique structural features of the pacidamycins. Heterologous expression in Streptomyces lividans resulted in the production of pacidamycin D and the newly identified pacidamycin S, thus confirming the identity of the pacidamycin biosynthetic gene cluster. Identification of this cluster will enable the generation of new uridyl peptide antibiotics through combinatorial biosynthesis. The concise cluster will provide a useful model system through which to gain a fundamental understanding of the way in which nonribosomal peptide synthetases interact.

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Lights on: a switchable fluorescent biliprotein.

Publication Date: 2010 Aug 16 PMID: 20665617
Authors: Gartner, W.
Journal: Chembiochem

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Molecular recognition of peloruside A by microtubules. The C24 primary alcohol is essential for biological activity.

Publication Date: 2010 Aug 16 PMID: 20665616
Authors: Pera, B. - Razzak, M. - Trigili, C. - Pineda, O. - Canales, A. - Buey, R. M. - Jimenez-Barbero, J. - Northcote, P. T. - Paterson, I. - Barasoain, I. - Diaz, J. F.
Journal: Chembiochem

Peloruside is a microtubule-stabilizing agent that targets the same site as laulimalide. It binds to microtubules with a 1:1 stoichiometry and with a binding affinity in the low-muM range; thereby reducing the number of microtubular protofilaments in the same way as paclitaxel. Although the binding affinity of the compound is comparable to that of the low-affinity stabilizing agent sarcodictyin, peloruside is more active in inducing microtubule assembly and is more cytotoxic to tumor cells; this suggests that the peloruside site is a more effective site for stabilizing microtubules. Acetylation of the C24 hydroxyl group results in inactive compounds. According to molecular modeling, this substitution at the C24 hydroxyl group presumably disrupts the interaction of the side chain with Arg320 in the putative binding site on alpha-tubulin. The binding epitope of peloruside on microtubules has been studied by using NMR spectroscopic techniques, and is compatible with the same binding site.

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A single molecular beacon probe is sufficient for the analysis of multiple nucleic acid sequences.

Publication Date: 2010 Aug 16 PMID: 20665615
Authors: Gerasimova, Y. V. - Hayson, A. - Ballantyne, J. - Kolpashchikov, D. M.
Journal: Chembiochem

Molecular beacon (MB) probes are dual-labeled hairpin-shaped oligodeoxyribonucleotides that are extensively used for real-time detection of specific RNA/DNA analytes. In the MB probe, the loop fragment is complementary to the analyte: therefore, a unique probe is required for the analysis of each new analyte sequence. The conjugation of an oligonucleotide with two dyes and subsequent purification procedures add to the cost of MB probes, thus reducing their application in multiplex formats. Here we demonstrate how one MB probe can be used for the analysis of an arbitrary nucleic acid. The approach takes advantage of two oligonucleotide adaptor strands, each of which contains a fragment complementary to the analyte and a fragment complementary to an MB probe. The presence of the analyte leads to association of MB probe and the two DNA strands in quadripartite complex. The MB probe fluorescently reports the formation of this complex. In this design, the MB does not bind the analyte directly; therefore, the MB sequence is independent of the analyte. In this study one universal MB probe was used to genotype three human polymorphic sites. This approach promises to reduce the cost of multiplex real-time assays and improve the accuracy of single-nucleotide polymorphism genotyping.

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RAF kinase inhibitors in cancer treatment: like a bull in a China shop?

Publication Date: 2010 Aug 16 PMID: 20648512
Authors: Robubi, A. - Waldmann, H. - Rauh, D.
Journal: Chembiochem

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Rational design of novel peptidic DnaK ligands.

Publication Date: 2010 Aug 16 PMID: 20648511
Authors: Liebscher, M. - Haupt, K. - Yu, C. - Jahreis, G. - Lucke, C. - Schiene-Fischer, C.
Journal: Chembiochem

The hsp70 chaperone DnaK from E. coli plays a major role in cellular stress response and is involved in assisted protein folding in vivo. By screening a combinatorial peptide library, we identified several DnaK-specific peptide ligands with nanomolar affinities, which are able to inhibit the secondary amide peptide bond cis/trans isomerase (APIase) activity of DnaK, as well as DnaK/DnaJ/GrpE-assisted refolding of firefly luciferase. Our designed DnaK inhibitors have the capability to penetrate E. coli cells and feature a high protease resistance. Once inside the cell, they physically target DnaK. NMR-based (1)H/(15)N-HSQC experiments furthermore confirmed that the designed peptidic ligands all bind in an identical manner to the conventional peptide-binding site of DnaK. The subsequent blocking of DnaK function apparently results in the observed antibacterial effects on E. coli cells, with minimum inhibitory concentrations in the range of 100 microM.

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Site-specific cytosine to uracil transition by using reversible DNA photo-crosslinking.

Publication Date: 2010 Aug 16 PMID: 20632434
Authors: Fujimoto, K. - Konishi-Hiratsuka, K. - Sakamoto, T. - Yoshimura, Y.
Journal: Chembiochem

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How to spoil the taste of insect prey? A novel feeding deterrent against ants released by larvae of the alder leaf beetle, Agelastica alni.

Publication Date: 2010 Aug 16 PMID: 20632433
Authors: Hilker, M. - Haberlein, C. - Trauer, U. - Bunnige, M. - Vicentini, M. O. - Schulz, S.
Journal: Chembiochem

Chemical defense of leaf beetle larvae (Chrysomelidae) against enemies is provided by secretions containing a wide range of deterrent compounds or by unpalatable hemolymph constituents. Here we report a new, very strong feeding deterrent against ants released by larvae of the alder leaf beetle Agelastica alni when attacked. The larvae release a defensive fluid from openings of pairwise, dorsolaterally located tubercles on the first to the eighth abdominal segments. The fluid, consisting of hemolymph and probably a glandular cell secretion, has previously been shown to contain a very stable, non-volatile feeding deterrent. The major deterrent component was isolated by repeated HPLC separation and analyzed by NMR and MS. The compound proved to be gamma-L-glutamyl-L-2-furylalanine (1), a novel dipeptide containing the unusual amino acid L-2-furylalanine. This amino acid, although synthetically well known, has not previously been reported from natural sources. The absolute configuration of the natural compound was elucidated by enantioselective gas chromatography after derivatization. The structure of the dipeptide was verified by the synthesis of several isomeric dipeptides. In bioassays a concentration of 1 microg microL(-1) was sufficient to deter polyphagous Myrmica rubra ants from feeding.

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A genetic circuit system based on quorum sensing signaling for directed evolution of quorum-quenching enzymes.

Publication Date: 2010 Aug 16 PMID: 20632431
Authors: Kim, J. H. - Lee, S. C. - Kyeong, H. H. - Kim, H. S.
Journal: Chembiochem

Quorum sensing is a cell-cell communication mechanism that is involved in the regulation of biological functions such as luminescence, virulence, and biofilm formation. Quorum-quenching enzymes, which interrupt quorum-sensing signaling through degradation of quorum-sensing molecules, have emerged as a new approach to controlling and preventing bacterial virulence and pathogenesis. In an effort to develop quorum-quenching enzymes with improved catalytic activities, a genetic circuit system based on acylhomoserine-lactone (AHL)-mediated quorum-sensing signaling was constructed. The genetic circuit system was composed of lux-R, lux-I promoter, beta-lactamase, and beta-lactamase inhibitor, and designed to confer antibiotic resistance on host cells expressing an AHL-degrading enzyme, thereby enabling rapid screening of quorum-quenching enzymes. To demonstrate the utility of the genetic circuit system, we attempted the directed evolution of the AHL hydrolase from Bacillus sp. The genetic circuit system was shown to be effective in screening of quorum-quenching enzymes with high catalytic efficiency. From these results it is expected that the genetic circuit system can be widely used for the isolation and directed evolution of quorum-quenching enzymes with greater potential.

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A chemoenzymatic route to diversify aminoglycosides enables a microarray-based method to probe acetyltransferase activity.

Publication Date: 2010 Aug 16 PMID: 20629012
Authors: Tsitovich, P. B. - Pushechnikov, A. - French, J. M. - Disney, M. D.
Journal: Chembiochem

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Design of lanthanide fingers: compact lanthanide-binding metalloproteins.

Publication Date: 2010 Aug 16 PMID: 20623571
Authors: am Ende, C. W. - Meng, H. Y. - Ye, M. - Pandey, A. K. - Zondlo, N. J.
Journal: Chembiochem

Lanthanides have interesting chemical properties; these include luminescent, magnetic, and catalytic functions. Toward the development of proteins incorporating novel functions, we have designed a new lanthanide-binding motif, lanthanide fingers. These were designed based on the Zif268 zinc finger, which exhibits a beta beta alpha structural motif. Lanthanide fingers utilize an Asp(2)Glu(2) metal-coordination environment to bind lanthanides through a tetracarboxylate peptide ligand. The iterative design of a general lanthanide-binding peptide incorporated the following key elements: 1) residues with high alpha-helix and beta-sheet propensities in the respective secondary structures; 2) an optimized big box alpha-helix N-cap; 3) a Schellman alpha-helix C-cap motif; and 4) an optional D-Pro-Ser type II' beta-turn in the beta-hairpin. The peptides were characterized for lanthanide binding by circular dichroism (CD), NMR, and fluorescence spectroscopy. In all instances, stabilization of the peptide secondary structures resulted in an increase in metal affinity. The optimized protein design was a 25-residue peptide that was a general lanthanide-binding motif; this binds all lanthanides examined in a competitive aqueous environment, with a dissociation constant of 9.3 microM for binding Er(3+). CD spectra of the peptide-lanthanide complexes are similar to those of zinc fingers and other beta beta alpha proteins. Metal binding involves residues from the N-terminal beta-hairpin and the C terminal alpha-helical segments of the peptide. NMR data indicated that metal binding induced a global change in the peptide structure. The D-Pro-Ser type II' beta-turn motif could be replaced by Thr-Ile to generate genetically encodable lanthanide fingers. Replacement of the central Phe with Trp generated genetically encodable lanthanide fingers that exhibited terbium luminescence greater than that of an EF-hand peptide.

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Two-step labeling of endogenous enzymatic activities by Diels-Alder ligation.

Publication Date: 2010 Aug 16 PMID: 20623570
Authors: Willems, L. I. - Verdoes, M. - Florea, B. I. - van der Marel, G. A. - Overkleeft, H. S.
Journal: Chembiochem

A ligation strategy based on the Diels-Alder [4+2] cycloaddition for the two-step activity-based labeling of endogenously expressed enzymes in complex biological samples has been developed. A panel of four diene-derivatized proteasome probes was synthesized, along with a dienophile-functionalized BODIPY(TMR) tag. These probes were applied in a Diels-Alder labeling procedure that enabled us to label active proteasome beta-subunits selectively in cellular extracts and in living cells. We were also able to label the activity of cysteine proteases in cell extracts by utilizing a diene-derivatized cathepsin probe. Importantly, the Diels-Alder strategy described here is fully orthogonal with respect to the Staudinger-Bertozzi ligation, as demonstrated by the independent labeling of different proteolytic activities by the two methods in a single experiment.

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In vivo efficacy of natural product-inspired irreversible kinase inhibitors.

Publication Date: 2010 Aug 16 PMID: 20623569
Authors: Barluenga, S. - Jogireddy, R. - Koripelly, G. K. - Winssinger, N.
Journal: Chembiochem

Hypothemycin and related resorcylic acid lactones (RAL) bearing a cis-enone moiety have emerged as an alternative pharmacophore to heterocyclic motifs for kinase inhibition, and are endowed with a unique selectivity filter based on the irreversible reaction with a subset of the kinome bearing a suitably positioned cysteine residue. Two prototypical examples of "edited" RAL were evaluated for antitumoral, antimetastatic and antiangiogenic efficacy in an orthotopic murine renal cell carcinoma (RENCA) model. Both compounds (3 and 5) are good inhibitors of VEGFRs in vitro, and inhibited tumor growth in vivo with comparable efficacy to sunitinib, an FDA-approved VEGFRs inhibitor. Compound 3 promoted lung metastasis to a similar extent as sunitinib, while compound 5 strongly inhibited lung metastasis. This study attests to the potential of irreversible kinase inhibitors and molecular editing of natural pharmacophores and provides encouraging results to a clinically significant problem.

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Metal-stimulated regulation of transcription by an artificial zinc-finger protein.

Publication Date: 2010 Aug 16 PMID: 20607778
Authors: Imanishi, M. - Nakaya, T. - Morisaki, T. - Noshiro, D. - Futaki, S. - Sugiura, Y.
Journal: Chembiochem

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A single-electrode, dual-potential ferrocene-PNA biosensor for the detection of DNA.

Publication Date: 2010 Aug 16 PMID: 20602405
Authors: Husken, N. - Gebala, M. - Schuhmann, W. - Metzler-Nolte, N.
Journal: Chembiochem

A Fc-PNA biosensor (Fc: ferrocenyl, C(10)H(9)Fe) was designed by using two electrochemically distinguishable recognition elements with different molecular information at a single electrode. Two Fc-PNA capture probes were therefore synthesized by N-terminal labeling different dodecamer PNA sequences with different ferrocene derivatives by click chemistry. Each of the two strands was thereby tethered with one specific ferrocene derivative. The two capture probes revealed quasi-reversible redox processes of the Fc(0/+) redox couple with a significant difference in their electrochemical half-wave potentials of Delta E(1/2)=160 mV. A carefully designed biosensor interface, consisting of a ternary self-assembled monolayer (SAM) of the two C-terminal cysteine-tethered Fc-PNA capture probes and 6-mercaptohexanol, was electrochemically investigated by square wave (SWV) and cyclic voltammetry (CV). The biosensor properties of this interface were analyzed by studying the interaction with DNA sequences that were complementary to either of the two capture probes by SWV. Based on distinct changes in both peak current and potential, a parallel identification of these two DNA sequences was successful with one interface design. Moreover, the primary electrochemical response could be converted by a simple mathematical analysis into a clear-cut electrochemical signal about the hybridization event. The discrimination of single-nucleotide polymorphism (SNP) was proven with a chosen single-mismatch DNA sequence. Furthermore, experiments with crude bacterial RNA confirm the principal suitability of this dual-potential sensor under real-life conditions.

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Effects of hapten density on the induced antibody repertoire.

Publication Date: 2010 Aug 16 PMID: 20602400
Authors: Li, Q. - Rodriguez, L. G. - Farnsworth, D. F. - Gildersleeve, J. C.
Journal: Chembiochem

Small peptides and oligosaccharides are important antigens for the development of vaccines and the production of monoclonal antibodies. Because of their small size, peptides and oligosaccharides are non-immunogenic on their own and typically must be conjugated to a larger carrier protein to elicit an immune response. Selection of a suitable carrier protein, conjugation method, and hapten density are critical for generating an optimal immune response. We used a glycan array to compare the repertoire of antibodies induced after immunizing with either low or high-density conjugates of the tumor-associated Tn antigen. At high hapten density, a broader range of antibodies was induced, and reactivity to the clustered Tn antigen was observed. In contrast, antibodies induced by the low-density conjugate had narrower reactivity and did not bind the clustered Tn antigen.

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The alpha/beta-hydrolase fold 3DM database (ABHDB) as a tool for protein engineering.

Publication Date: 2010 Aug 16 PMID: 20593436
Authors: Kourist, R. - Jochens, H. - Bartsch, S. - Kuipers, R. - Padhi, S. K. - Gall, M. - Bottcher, D. - Joosten, H. J. - Bornscheuer, U. T.
Journal: Chembiochem

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Protease-activatable collagen targeting based on protein cyclization.

Publication Date: 2010 Aug 16 PMID: 20589824
Authors: Breurken, M. - Lempens, E. H. - Merkx, M.
Journal: Chembiochem

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