Use of a thiol tether for the site-specific attachment of reporter groups to DNA

Fidanza, Jacqueline A.; McLaughlin, Larry W.

doi:10.1021/jo00034a028

Cited by 45 publications

(26 citation statements)

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“…[5] We used two DNA sequences, 5'-CGGCTpACTCC-3' (1 XL) and 5'-GTGCTpAGCGG-3' (2 XL; p denotes a phosphoramidate linkage where a cross-linker is introduced). Analysis by reversed-phase HPLC revealed two diastereomers, A (faster eluting) and B (slower eluting; see Supporting Information).…”

Section: Masayuki Endo and Tetsuro Majima*mentioning

confidence: 99%

“…The length of the bismaleimide linkers affected the stabilities of the DNA assemblies, and the crosslinked oligonucleotides with a C 2 linker formed more-stable assemblies than those with a C 6 linker. The stereochemical effects of the phosphoramidates were also observed, [5] and the assemblies containing the B diastereomers were thermally more stable than those with the A diastereomers. …”

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confidence: 95%

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Parallel, Double‐Helix DNA Nanostructures Using Interstrand Cross‐Linked Oligonucleotides with Bismaleimide Linkers

Endo¹,

Majima²

2003

Angew Chem Int Ed

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The creation of functional nanometer-scale wires and arrays has attracted increasing interest in the field of material sciences and molecular electronics.[1] For this purpose, molecular scaffolds, including polymers and supramolecules, with well-defined architectures are required for placing and arranging the desired functional molecules.[1] Nucleic acids provide a good scaffold for the following reasons: 1) The structure of DNA is well-defined and considered to be a periodic double-helical polymer; 2) association of DNA strands can be controlled by programming the sequence of DNA; 3) DNA molecules are easily modified chemically and biochemically for introduction of various functional molecules and nanoparticles; [2] and 4) DNA can be manipulated with modifying enzymes such as DNA ligase, kinase, polymerase, and restriction enzymes. Although DNA is an ideal scaffold, the flexibility of a long DNA chain makes the construction of desirable rigid structures difficult. Seeman and co-workers have built micrometer-scale DNA architectures by arranging previously prepared rigid DNA components.[3] The preparation and creation of wires and arrays would be much easier if such rigid DNA structures could be prepared simply by the addition of a chemical connector to the DNA strands.Here we describe the design, synthesis, and properties of novel cross-linked oligonucleotides (XL-DNA) for the construction of a micrometer-scale rodlike DNA structure. Crosslinked oligonucleotides were designed to connect two doublehelix DNA strands using a tether from a diastereochemically pure phosphoramidate (Scheme 1). Interstrand DNA crosslinking has been investigated in previous studies in which modification of a specific nucleic acid base was employed. [4] Our design is advantageous for connecting two duplex strands with less structural stress because the phosphate groups are located in the outer positions of the double-helical DNA. We employed a bifunctional cross-linker, alkyl bismaleimide, which can form a covalent bond with a thiol residue of a DNA strand under mild conditions. In addition, the distance between two DNA strands can be controlled using two different lengths of bismaleimide linkers.The diastereochemically pure oligonucleotides were prepared according to a literature method.[5] We used two DNA sequences, 5'-CGGCTpACTCC-3' (1 XL) and 5'-GTGCTpAGCGG-3' (2 XL; p denotes a phosphoramidate linkage where a cross-linker is introduced). Analysis by reversed-phase HPLC revealed two diastereomers, A (faster eluting) and B (slower eluting; see Supporting Information). The DNA was cross-linked according to Scheme 2. Oligonucleotides having a cystamine linker were treated with dithiothreitol (DTT) to reduce the disulfide linker. Thioltethered DNA (SH-DNA) was treated with excess 1,2-bismaleimidoethane (C 2 ) or 1,6-bismaleimidohexane (C 6 ) to produce a monocapped bismaleimide-DNA conjugate. This DNA conjugate and the SH-DNA were incubated in a 1:1 mol ratio, and the resulting XL-DNA was purified by HPLC. The extent of cross-linking w...

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Section: Masayuki Endo and Tetsuro Majima*mentioning

confidence: 99%

mentioning

confidence: 95%

Parallel, Double‐Helix DNA Nanostructures Using Interstrand Cross‐Linked Oligonucleotides with Bismaleimide Linkers

Endo¹,

Majima²

2003

Angew Chem Int Ed

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“…A variety of reporter groups have been attached sequence-specifically to oligodeoxynucleotides by the oxidation of an internucleotidic H-phosphonate in presence of cystamine to form a phosphoramidate linkage (242). Reduction of cystamine results in a free sulfhydryl group that can act as a nucleophile to react with thiol-specific reporter groups.…”

Section: Fluorescent Labelsmentioning

confidence: 99%

“…This derivative seems to be particularly attractive because the motion of the spin label can be directly correlated to that of the base. A nitroxide spin label has also been introduced at the phosphate backbone, via a phosphoramidate sulfhydryl tether, but EPR measurements were not reported (242).…”

Section: Electron Spin Resonance Probesmentioning

confidence: 99%

MODIFIED OLIGONUCLEOTIDES: Synthesis and Strategy for Users

Verma

Eckstein²

1998

Annu. Rev. Biochem.

299

190

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Synthetic oligonucleotide analogs have greatly aided our understanding of several biochemical processes. Efficient solid-phase and enzyme-assisted synthetic methods and the availability of modified base analogs have added to the utility of such oligonucleotides. In this review, we discuss the applications of synthetic oligonucleotides that contain backbone, base, and sugar modifications to investigate the mechanism and stereochemical aspects of biochemical reactions. We also discuss interference mapping of nucleic acid-protein interactions; spectroscopic analysis of biochemical reactions and nucleic acid structures; and nucleic acid cross-linking studies.The automation of oligonucleotide synthesis, the development of versatile phosphoramidite reagents, and efficient scale-up have expanded the application of modified oligonucleotides to diverse areas of fundamental and applied biological research. Numerous reports have covered oligonucleotides for which modifications have been made of the phosphodiester backbone, of the purine and pyrimidine heterocyclic bases, and of the sugar moiety; these modifications serve as structural and mechanistic probes. In this chapter, we review the range, scope, and practical utility of such chemically modified oligonucleotides. Because of space limitations, we discuss only those oligonucleotides that contain phosphate and phosphate analogs as internucleotidic linkages. CONTENTS

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“…McLaughlin and co-workers have incorporated a thiol into the sugar-phosphate backbone of DNA by oxidation of an internucleotide H-phosphonate at a unique site in the presence of cystamine to produce an internal phosphoramidate (Fig. 2b) (12). This approach has the same disadvantages as alkylation of phosphorothioates, namely, that a diastereomeric mixture is obtained which also contains a rather unstable N-substituted phosphoramidate.…”

Section: Academic Pressmentioning

confidence: 99%

Thiol-Containing RNA for the Study of Structure and Function of Ribozymes

Sigurdsson

1999

Methods

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Use of a thiol tether for the site-specific attachment of reporter groups to DNA

Cited by 45 publications

References 0 publications

Parallel, Double‐Helix DNA Nanostructures Using Interstrand Cross‐Linked Oligonucleotides with Bismaleimide Linkers

Parallel, Double‐Helix DNA Nanostructures Using Interstrand Cross‐Linked Oligonucleotides with Bismaleimide Linkers

MODIFIED OLIGONUCLEOTIDES: Synthesis and Strategy for Users

Thiol-Containing RNA for the Study of Structure and Function of Ribozymes

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