Cover image for ChemBioChem

December 2, 2005

Volume 6, Issue 12

Pages 2113–2342

    1. Cover Picture: Sequence-Specific DNA Recognition by Monomeric bZIP Basic Regions Equipped with a Tripyrrole Unit on the N-Terminal Side. Towards the Development of Synthetic Mimics of Skn-1 (ChemBioChem 12/2005) (page 2113)

      Juan B. Blanco, M. Eugenio Vázquez, Luis Castedo and José L. Mascareñas

      Version of Record online: 30 NOV 2005 | DOI: 10.1002/cbic.200590039

      The cover picture shows a representation of a new DNA-recognition strategy based on clamping dsDNA by the simultaneous interaction of a tripyrrole and the basic region of transcription factor GCN4 with two contiguous DNA sites. The peptide unit by itself is incapable of tight specific DNA binding; however, conjugation with a tripyrrole derivative capable of binding to AT-rich sites allows the specific delivery of the hybrid to composite DNA sites of 8–9 base pairs containing the consensus target sequences of each module. For more details, see the communication by J. L. Mascareñas et al. on p. 2173 ff.

    2. Break on through to the Other Side—Biophysics and Cell Biology Shed Light on Cell-Penetrating Peptides (pages 2126–2142)

      Rainer Fischer, Mariola Fotin-Mleczek, Hansjörg Hufnagel and Roland Brock

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500044

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      Finding the way in. Despite the broad acceptance of cell-penetrating peptides (CPPs) as molecular carriers, the details of the mode of cellular internalization and membrane permeation of these molecules still remain elusive. A detailed mechanistic understanding of the import mechanism will depend on a close interaction between cell biology and biophysics.

    3. Allosteric Control of mRNA Decoding (pages 2143–2145)

      Mathias Sprinzl and C. Stefan Voertler

      Version of Record online: 25 OCT 2005 | DOI: 10.1002/cbic.200500299

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      Ribosomal protein biosynthesis is a chemical reaction with remarkable efficiency and fidelity. The chemical mechanisms enabling this remain unclear, despite all the structural insight gained over the years. Recent results on the effect of remote mutations give some clues: the entire tRNA conformation can overrule the codon–anticodon decoding (see figure).

    4. Selective Staudinger Modification of Proteins Containing p-Azidophenylalanine (pages 2147–2149)

      Meng-Lin Tsao, Feng Tian and Peter G. Schultz

      Version of Record online: 30 NOV 2005 | DOI: 10.1002/cbic.200500314

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      The unnatural amino acidp-azidophenylalanine was genetically introduced into a phage-displayed peptide and mutant Z-domain protein. These azide-containing proteins were selectively and efficiently modified by using fluorescein-derived phosphines and the Staudinger ligation reaction (see picture). This work provides a powerful new tool for the selective modification of proteins with a variety of synthetic agents.

    5. A DNA Mimic Made of Non-Nucleosidic Phenanthrene Building Blocks (pages 2149–2152)

      Simon M. Langenegger and Robert Häner

      Version of Record online: 4 NOV 2005 | DOI: 10.1002/cbic.200500366

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      Stable stacking. DNA mimics have been prepared from simple, non-nucleosidic building blocks. These oligonucleotides, which contain extended stretches of phenanthrene derivatives, form stable hybrids. The phenanthrene residues contribute to the stability of the duplex through interstrand stacking interactions. The hybrids adopt an overall B-form structure, thus mimicking natural DNA.

    6. Circular Dichroism of Designed Peptide Helices and β-Hairpins: Analysis of Trp- and Tyr-Rich Peptides (pages 2152–2158)

      Radhakrishnan Mahalakshmi, Ganesh Shanmugam, Prasad L. Polavarapu and Padmanabhan Balaram

      Version of Record online: 31 OCT 2005 | DOI: 10.1002/cbic.200500152

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      VCD versus ECD spectroscopy. Peptides rich in aromatic residues yield anomalous far-UV electronic circular dichroism (ECD) spectra that preclude secondary structure assignment. The utility of vibrational circular dichroism (VCD) in conformation analysis is demonstrated by using a set of well-defined peptide helices and hairpins containing proximal aromatic residues.

    7. Site-Specific Fluorescent and Affinity Labelling of RNA by Using a Small Engineered Twin Ribozyme (pages 2158–2162)

      Stéphanie Vauléon, Sergei A. Ivanov, Slawomir Gwiazda and Sabine Müller

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500215

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      Site-determined functionalization of large RNA molecules can be achieved by a strategy involving twin ribozymes and small synthetic oligonucleotides. The twin ribozyme mediates the exchange of a predefined patch of RNA sequence against a modified oligonucleotide to result in specific functionalization of the large RNA.

    8. Quantum-Dot Aptamer Beacons for the Detection of Proteins (pages 2163–2166)

      Matthew Levy, Sean F. Cater and Andrew D. Ellington

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500218

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      Seeing thrombin in real time. Quantum-dot aptamer beacons provide for the sensitive detection of protein targets, such as thrombin, as well as nucleic acid targets and offer the promise of simplified, multiplex detection.

    9. Evidence for the Unusual Condensation of a Diketide with a Pentulose in the Methylenomycin Biosynthetic Pathway of Streptomyces coelicolor A3(2) (pages 2166–2170)

      Christophe Corre and Gregory L. Challis

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500243

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      How is the extraordinarily dense array of functional groups adorning the cyclopentane rings of the antibiotic methylenomycins assembled? Here it is shown that the northern 5 carbons of the methylenomycins (blue) derive from a pent(ul)ose intermediate of the pentose phosphate pathway. Together with previous results showing that the southern 4 carbons (green) derive from two acetate units and analysis of the mmy gene cluster that directs methylenomycin biosynthesis in Streptomyces coelicolor A3(2), these results suggest plausible early steps for methylenomycin biosynthesis via a butenolide intermediate.

    10. Biosynthetic Incorporation of Fluorohistidine into Proteins in E. coli: A New Probe of Macromolecular Structure (pages 2170–2173)

      Jack F. Eichler, John C. Cramer, Kenneth L. Kirk and James G. Bann

      Version of Record online: 31 OCT 2005 | DOI: 10.1002/cbic.200500249

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      Fluorine, the other hydrogen: Fluorinated forms of histidine, namely 2- and 4-fluorohistidines (see scheme), have dramatically reduced side-chain pKa's compared to the native amino acid. Biosynthetic incorporation of these analogues into bacterially expressed proteins should, therefore, broaden studies that are aimed at understanding pH- dependent processes. In this study we apply this technique to the production of fluorinated PapD chaperone.

    11. Sequence-Specific DNA Recognition by Monomeric bZIP Basic Regions Equipped with a Tripyrrole Unit on the N-Terminal Side. Towards the Development of Synthetic Mimics of Skn-1 (pages 2173–2176)

      Juan B. Blanco, M. Eugenio Vázquez, Luis Castedo and José L. Mascareñas

      Version of Record online: 17 OCT 2005 | DOI: 10.1002/cbic.200500294

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      Appropriate conjugation of distamycin-related tripyrroles with an N-terminal position of a peptide containing the basic region amino acids of a bZIP protein (see figure) provide for high affinity recognition of composite 8–9 bp DNA sites.

    12. Tetrabutylammonium Fluoride-Mediated Rapid Alkylation Reaction in Microtiter Plates for the Discovery of Enzyme Inhibitors in Situ (pages 2176–2180)

      Chung-Yi Wu, Ashraf Brik, Sheng-Kai Wang, Yu-Hsien Chen and Chi-Huey Wong

      Version of Record online: 4 NOV 2005 | DOI: 10.1002/cbic.200500295

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      No isolation, no purification. An efficient and simple method based on tetrabutylammonium fluoride-mediated alkylation without protecting groups in microtiter plates, followed by screening in situ has been developed to identify new enzyme inhibitors, as demonstrated in the discovery of the shown potent inhibitors of cathepsin B and HIV protease.

    13. Ligand Binding Transmits Conformational Changes across the Membrane-Spanning Region to the Intracellular Side of the 5-HT3 Serotonin Receptor (pages 2180–2185)

      Erwin Ilegems, Horst Pick, Cédric Deluz, Stephan Kellenberger and Horst Vogel

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500191

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      Conformational changes of channel activation: Five enhanced green fluorescent protein (EGFP) molecules (green cylinders) were integrated into the intracellular part of the homopentameric ionotropic 5-HT3 receptor. This allowed the detection of extracellular binding of fluorescent ligands (•) to EGFP by FRET, and also enabled the quantification of agonist-induced conformational changes in the intracellular region of the receptor by homo-FRET between EGFPs. The approach opens novel ways for probing receptor activation and functional screening of therapeutic compounds.

    14. Reversible Sequential-Binding Probe Receptor–Ligand Interactions in Single Cells (pages 2187–2194)

      Christoph Schreiter, Marinela Gjoni, Ruud Hovius, Karen L. Martinez, Jean-Manuel Segura and Horst Vogel

      Version of Record online: 4 NOV 2005 | DOI: 10.1002/cbic.200500216

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      Time after time. A new method called the reversible sequential (ReSeq)-binding assay is presented. We demonstrate the application of this technique to the ultrasensitive profiling of receptor function in single living cells. The assay is based on the repetitive application of fluorescent ligands that have fast association–dissociation kinetics (see figure). This system could become an important tool for functional screening of lead compounds.

    15. Discovery of Complex Mixtures of Novel Long-Chain Quorum Sensing Signals in Free-Living and Host-Associated Marine Alphaproteobacteria (pages 2195–2206)

      Irene Wagner-Döbler, Verena Thiel, Leo Eberl, Martin Allgaier, Agnes Bodor, Sandra Meyer, Sabrina Ebner, Andreas Hennig, Rüdiger Pukall and Stefan Schulz

      Version of Record online: 11 NOV 2005 | DOI: 10.1002/cbic.200500189

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      Cell-density-dependent regulation of gene expression, or quorum sensing, enables bacteria to coordinate their behaviour and plays an important role during infection. Using a new method to extract acylated homoserine lactone (AHL) quorum-sensing signals from culture media and combining a highly sensitive biological detection method with nontarget chemical screening, we discovered novel long-chain AHLs in more than half of the marine Alphaproteobacteria studied, such as the aerobic anoxygenic phototroph Dinoroseobacter shibae is shown here.

    16. Dendritic Catanionic Assemblies: In vitro Anti-HIV Activity of Phosphorus-Containing Dendrimers Bearing Galβ1cer Analogues (pages 2207–2213)

      Muriel Blanzat, Cédric-Olivier Turrin, Anne-Marie Aubertin, Christiane Couturier-Vidal, Anne-Marie Caminade, Jean-Pierre Majoral, Isabelle Rico-Lattes and Armand Lattes

      Version of Record online: 30 NOV 2005 | DOI: 10.1002/cbic.200500203

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      Biological activity of galactosylceramide analogues: A series of water-soluble catanionic phosphorus-containing low-generation dendritic assemblies mimicking multisite analogues of galactosylceramide exhibit structure-dependent in vitro anti-HIV-1 activity. The shape-dependent activity is modulated by a dendritic multivalency effect.

    17. Fluorescent Detection of Apoptotic Cells by Using Zinc Coordination Complexes with a Selective Affinity for Membrane Surfaces Enriched with Phosphatidylserine (pages 2214–2220)

      Roger G. Hanshaw, C. Lakshmi, Timothy N. Lambert, James R. Johnson and Bradley D. Smith

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500149

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      Going out with a glow. The appearance of phosphatidylserine on the membrane surface is a hallmark of the process of programmed cell death (also known as apoptosis). A series of chemical agents are shown to selectively stain the surface of apoptotic cells; thus, they act as low-molecular-weight mimics of the membrane binding protein annexin V.

    18. Access of the Substrate to the Active Site of Yeast Oxidosqualene Cyclase: An Inhibition and Site-Directed Mutagenesis Approach (pages 2221–2228)

      Simonetta Oliaro-Bosso, Tanja Schulz-Gasch, Gianni Balliano and Franca Viola

      Version of Record online: 19 OCT 2005 | DOI: 10.1002/cbic.200500107

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      Holding it all together: Modeling, inhibition, and site-directed mutagenesis suggest the existence in yeast oxidosqualene cyclase of a channel constriction that permits the access of the substrate to the active site. A single residue of the constriction loop, Glu526 (see model), appears to be a keystone of the whole architecture of the protein.

    19. Carbohydrate Array Analysis of Anti-Tn Antibodies and Lectins Reveals Unexpected Specificities: Implications for Diagnostic and Vaccine Development (pages 2229–2241)

      Joseph C. Manimala, Zhitao Li, Amit Jain, Sharanjeet VedBrat and Jeffrey C. Gildersleeve

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500165

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      A case of mistaken identity. A carbohydrate microarray containing a variety of epitopes such as tumor antigens, blood-group antigens, and Lewis antigens was developed and used to evaluate the specificity of several anti-Tn antibodies and lectins. Array analysis revealed unexpected cross-reactivity with several human carbohydrates; this indicates that other epitopes can be mistaken for the tumor-associated Tn antigen.

    20. Solution NMR Studies of an Intrinsically Unstructured Protein within a Dilute, 75 kDa Eukaryotic Protein Assembly; Probing the Practical Limits for Efficiently Assigning Polypeptide Backbone Resonances (pages 2242–2246)

      Yuefeng Wang, Igor Filippov, Christian Richter, Rensheng Luo and Richard W. Kriwacki

      Version of Record online: 4 NOV 2005 | DOI: 10.1002/cbic.200500260

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      Cold feat. A combined strategy to obtain the backbone assignments of an intrinsically unstructured protein, p21-KID, bound to its biological target, Cdk2/cyclin A, is presented. In order to overcome the challenges associated with the high molecular weight and low solubility of the ternary complex, we used perdeuteration, specific amino acid isotope labeling, TROSY, and cryogenic NMR probes at high magnetic-field strengths. In addition, we studied binary subcomplexes; this allowed resonance assignments to be made in stages.

    21. Binding of Helix-Threading Peptides to E. coli 16S Ribosomal RNA and Inhibition of the S15–16S Complex (pages 2247–2254)

      Barry D. Gooch, Malathy Krishnamurthy, Mohammad Shadid and Peter A. Beal

      Version of Record online: 24 OCT 2005 | DOI: 10.1002/cbic.200500285

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      Catch 22. Helix-threading peptides (HTPs) that bind helix 22 of E. coli 16S RNA and block interaction with the ribosomal protein S15 are described. HTP binding is dependent on the presence of a highly conserved, purine-rich internal loop that influences conformational flexibility at the binding site.

    22. Design, Synthesis and Analysis of Inhibitors of Bacterial Aspartate Semialdehyde Dehydrogenase (pages 2255–2260)

      Russell J. Cox, Jennifer S. Gibson and Andrea T. Hadfield

      Version of Record online: 31 OCT 2005 | DOI: 10.1002/cbic.200500172

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      Docking fluorophosphonates. Simulated docking of unsaturated and fluorinated phosphonates into the active site of bacterial aspartate semialdehyde dehydrogenase reveals binding interactions that explain the inhibitory activity of synthetic compounds.

    23. Protein Flexibility and Ligand Rigidity: A Thermodynamic and Kinetic Study of ITAM-Based Ligand Binding to Syk Tandem SH2 (pages 2261–2270)

      Nico J. de Mol, M. Isabel Catalina, Frank J. Dekker, Marcel J. E. Fischer, Albert J. R. Heck and Rob M. J. Liskamp

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500141

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      High affinity and fast dissociation kinetics. Protein dynamics might have strong implications for ligand design. Here it is shown by thermodynamic and kinetic analysis, as well as by protein-dynamics studies, how the flexibility of the receptor affects the binding of a flexible and rigid ligand to the Syk tandem SH2 domain. A bivalent binding model that combines high affinity with rapid dissociation kinetics is proposed. Both are relevant in transient signal-transduction processes.

    24. Development of a Novel Chemical Probe for the Selective Enrichment of Phosphorylated Serine- and Threonine-Containing Peptides (pages 2271–2280)

      Pieter van der Veken, Eef H. C. Dirksen, Eelco Ruijter, Ronald C. Elgersma, Albert J. R. Heck, Dirk T. S. Rijkers, Monique Slijper and Rob M. J. Liskamp

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500209

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      A multifunctional probe (shown) for the selective enrichment of serine- and threonine-phosphorylated peptides has been developed. Phosphopeptides are modified to allow probe attachment and affinity enrichment, facilitating MS analysis after removal of the affinity tag.

    25. Design and Synthesis of Oligosaccharides that Interfere with Glycoprotein Quality-control systems (pages 2281–2289)

      Midori A. Arai, Ichiro Matsuo, Shinya Hagihara, Kiichiro Totani, Jun-ichi Maruyama, Katsuhiko Kitamoto and Yukishige Ito

      Version of Record online: 11 NOV 2005 | DOI: 10.1002/cbic.200500143

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      Calnexin (CNX) and calreticulin (CRT), lectin-like molecular chaperones: Together with glucosidase II (Glc-ase II) and glycosyltransferase (UGGT) they constitute the elements of the CNX/CRT cycle, which assists the folding of newly synthesized glycoproteins. A non-natural undecasaccaride (1) and a heptasaccharide (2) were designed and synthesized as potential inhibitors of the ER quality-control system.

    26. Pityriarubins, Novel Highly Selective Inhibitors of Respiratory Burst from Cultures of the Yeast Malassezia furfur: Comparison with the Bisindolylmaleimide Arcyriarubin A (pages 2290–2297)

      Hans-Joachim Krämer, Dino Kessler, Ute-Christina Hipler, Bernhard Irlinger, Wiebke Hort, Rolf-Hasso Bödeker, Wolfgang Steglich and Peter Mayser

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500163

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      Highly selective inhibitors of ROS release in granulocytes. The bright red pityriarubins A (shown), B and C, isolated from cultures of the yeast Malassezia furfur in presence of tryptophan, structurally resemble the potent protein kinase inhibitors of the aryriarubin A family. They exhibited remarkable selectivity in suppressing granulocytary ROS release with respect to various agents.

    27. Synthesis and Properties of Aminopropyl Nucleic Acids (pages 2298–2304)

      Ding Zhou, Irene M. Lagoja, Jef Rozenski, Roger Busson, Arthur Van Aerschot and Piet Herdewijn

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500170

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      Great pretenders. When incorporated into DNA, aminopropyl nucleic acids (APNAs) do not show favourable hybridization properties in a mixed sequence. Here we show that the (S)-APNA series (see scheme) is slightly better accommodated and that an aminopropyl adenosine pairing (A*–A*) is especially favourable. In addition (R)-A* behaves almost like the best ambiguous nucleoside analogues known to-date and deserves further study.

    28. Design of a Hairpin Polyamide, ZT65B, for Targeting the Inverted CCAAT Box (ICB) Site in the Multidrug Resistant (MDR1) Gene (pages 2305–2311)

      Karen L. Buchmueller, Zarmeen Taherbhai, Cameron M. Howard, Suzanna L. Bailey, Binh Nguyen, Caroline O’Hare, Daniel Hochhauser, John A. Hartley, W. David Wilson and Moses Lee

      Version of Record online: 27 OCT 2005 | DOI: 10.1002/cbic.200500179

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      Out of the box: A novel 3-methylpicolinate-containing hairpin polyamide, ZT65B, has been designed and synthesized. Results from DNase I footprinting, surface-plasmon resonance, circular dichroism, and DNA thermal-melt experiments provided evidence for ZT65B's specific binding to the 3′ site (TGGCT) of the inverted CCAAT-box sequence found within the MDR1 promoter.

    29. LanV, a Bifunctional Enzyme: Aromatase and Ketoreductase during Landomycin A Biosynthesis (pages 2312–2315)

      Almuth Mayer, Takaaki Taguchi, Anton Linnenbrink, Carsten Hofmann, Andriy Luzhetskyy and Andreas Bechthold

      Version of Record online: 11 NOV 2005 | DOI: 10.1002/cbic.200500205

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      Jack of all trades. The exact function of LanV, which is involved in the biosynthesis of landomycin A (see scheme), was elucidated by combinatorial biosynthesis. Our results show that LanV is involved in the 6-ketoreduction of the angucycline core and catalyses the aromatization of ring A of the angucycline structure.

    30. Acyl Transfer in Clorobiocin Biosynthesis: Involvement of Several Proteins in the Transfer of the Pyrrole-2-carboxyl Moiety to the Deoxysugar (pages 2316–2325)

      Anja Freitag, Emmanuel Wemakor, Shu-Ming Li and Lutz Heide

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500252

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      Deletion ofcloN1andcloN7in the biosynthetic gene cluster of clorobiocin: The modified gene clusters were expressed in the heterologous host Streptomyces coelicolor M512 to give free pyrrole-2-carboxylic acid and a clorobiocin derivative that lacked the pyrrole-2-carboxyl moiety. These results showed that CloN1 and CloN7, together with the previously investigated CloN2, are involved in the transfer of the pyrrole-2-carboxyl moiety to the deoxysugar of clorobiocin.

    31. Use of Short Duplexes for the Analysis of the Sequence-Dependent Cleavage of DNA by a Chemical Nuclease, a Manganese Porphyrin (pages 2326–2335)

      Sophie Mourgues, Adam Kupan, Geneviève Pratviel and Bernard Meunier

      Version of Record online: 7 NOV 2005 | DOI: 10.1002/cbic.200500254

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      O-atom transfer versus electron transfer. Comparative oxidative DNA damage on different short DNA duplexes by the manganese(III)-bis(aqua)-meso-tetrakis(4-N-methylpyridiniumyl)porphyrin (see structure) in the presence of KHSO5 shows that guanine oxidation by electron transfer is not competitive with deoxyribose damage by O-atom transfer at (AT)3 sites on DNA, except for accessible guanines like the terminal guanine residue of a short duplex.

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      Version of Record online: 30 NOV 2005 | DOI: 10.1002/cbic.200590041