Sequenz‐spezifischer Einbau von Enzym‐Nukleotid‐Chimären durch DNA‐Polymerasen

Welter, Moritz; Verga, Daniela; Marx, Andreas

doi:10.1002/ange.201604641

“…17 In this context, Prof. Marx presented their explorations testing the potential of protein-modified nucleotides as a tool for diagnosing mutations by the naked eye using glycoprotein horseradish peroxidase (HPR) proteins. 18 This colorimetric method for the detection of nucleic acids provides a high fidelity readout, since only primers that bind to a specific target sequence will be modified by HRP due to the specificity of DNA polymerases.…”

mentioning

confidence: 99%

Highlights from the 53rd EUCHEM conference on stereochemistry, Bürgenstock, Switzerland, May 2018

Milo

¹

,

Temprano

²

2018

Chem. Commun.

0

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When we first heard of the Bürgenstock conference it was described as a guarded meeting in a remote location, undisturbed by modern diversions, with mysterious customs and a secret handshake. All of these rumors turned out to be completely true. We arrived at an undisclosed location and three young men, who knew our names upon sight, greeted us and gave us an agenda in which we discovered the identity of the speakers, moderators and other participants. We then had a few moments to bask in their glory before being treated to a delicious dose of chemistry. During the banquet, before the first lecture, this year's president, Prof. Ilan Marek, gave the opening address from a balcony reserved only for these meetings, rumors hold that it stands vacant all year in anticipation. He mentioned several of the traditions of the meeting, which we are not allowed to share of course, and described the joy of putting together this year's exciting program with the help of an exceptional organizing committee that included Prof. Cristina Nevado, Prof. Christian Bochet, Dr Fabrice Gallou and Dr Alain De Mesmaeker. He also introduced this year's guest of honor, Prof. Yitzhak Apeloig, and wished the best luck to the current Vice President and future President, Prof. Véronique Gouverneur, not only for preparing the 54th Bürgenstock Conference, but also with her duty this year, maintaining the good weather for the whole week. Although the official title of the conference is the EUCHEM conference on stereochemistry, the topics covered span a broad range of cutting edge chemical transformations and insights, which can appeal to anyone working in chemistry and its interfaces with other disciplines. We will briefly describe each of the talks following one of our favorite citable quotes from these inspiring speakers.

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2‐Substituted dATP Derivatives as Building Blocks for Polymerase‐Catalyzed Synthesis of DNA Modified in the Minor Groove

Matyašovský

¹

,

Perlíková

²

,

Malnuit

³

et al. 2016

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2'-Deoxyadenosine triphosphate (dATP) derivatives bearing diverse substituents (Cl, NH 2 ,CH 3 ,vinyl, ethynyl, and phenyl) at position 2were prepared and tested as substrates for DNApolymerases.The 2-phenyl-dATP was not asubstrate for DNApolymerases,but the dATPs bearing smaller substituents were good substrates in primer-extension experiments,producing DNAsubstituted in the minor groove. The vinyl-modified DNAw as applied in thiol-ene addition and the ethynylmodified DNAwas applied in aCuAACclick reaction to form DNAlabelled with fluorescent dyes in the minor groove Base-modified oligonucleotides (ONs) or DNAa re widely used as tools in chemical biology,d iagnostics,o rm aterials science.[1] Themodification is mostly attached to position 5of pyrimidines or position 7of7-deazapurines,not only because it then points out into the major groove of DNAand thus does not destabilize the duplex, but because in most cases,t he corresponding substituted 2'-deoxyribonucleoside triphosphates (dNTPs) are good substrates for DNAp olymerases and can be used in the polymerase-catalyzed synthesis of modified DNA. [2,3] Diverse modifications,i ncluding fluorophores, [4] redox [5] or spin labels, [6] reactive groups for conjugations, [7] and biomolecules (e.g., oligonucleotides [8] or proteins [9] ), have been introduced into the major groove through the enzymatic incorporation of modified nucleotides and applied in different fields.Modification or labelling of the minor groove has mostly been reported with 2'-and 4'-sugarmodified derivatives.[10-13] 2-Chloroadenine [14] and 2,6-diaminopurine [15] dNTPs are the only minor-groove base-modified nucleotides that have been reported as substrates for DNA polymerases,w hereas 2-arylamino-dATP derivatives were found to act as polymerase inhibitors.[16] Them inor groove sites of the nucleobases are difficult to modify since they are crucial both for Watson-Crick base pairing and for key minorgroove interactions with DNApolymerase that are important for extension of the chain.[17] On the other hand, 2-ethynylpyridone-C-nucleotide incorporated into DNA [18] formed as table base pair with adenine,a nd 2-(imidazolylalkylamino)purines in ONs also stabilized duplexes. [19] Because the possibility of minor-groove base labelling would be attractive for many prospective applications,for example,the mapping of DNA-protein interactions,w ee nvisaged that as mall substituent at position 2ofapurine may not fully disturb the key H-bonding interactions with the opposite base and the polymerase,a nd we report herein the first enzymatic synthesis of minor-groove base-modified DNA.Aseries of six 2-substituted dATP derivatives bearing Cl, NH 2 ,C H 3 ,v inyl, ethynyl and phenyl substituents (d R ATPs) was designed to study the effect of substituents of different bulkiness at position 2o fa denine on polymerase-mediated incorporation. While dCl ATP [14,20] and d NH2 ATP [15] were known, the others were prepared through triphosphorylation [21] of the corresponding 2'-deoxy-ribonucleosides (d R As...

show abstract

2‐Substituted dATP Derivatives as Building Blocks for Polymerase‐Catalyzed Synthesis of DNA Modified in the Minor Groove

Matyašovský

¹

,

Perlíková

²

,

Malnuit

³

et al. 2016

View full text Add to dashboard Cite

2'-Deoxyadenosine triphosphate (dATP) derivatives bearing diverse substituents (Cl, NH 2 ,CH 3 ,vinyl, ethynyl, and phenyl) at position 2were prepared and tested as substrates for DNApolymerases.The 2-phenyl-dATP was not asubstrate for DNApolymerases,but the dATPs bearing smaller substituents were good substrates in primer-extension experiments,producing DNAsubstituted in the minor groove. The vinyl-modified DNAw as applied in thiol-ene addition and the ethynylmodified DNAwas applied in aCuAACclick reaction to form DNAlabelled with fluorescent dyes in the minor groove Base-modified oligonucleotides (ONs) or DNAa re widely used as tools in chemical biology,d iagnostics,o rm aterials science.[1] Themodification is mostly attached to position 5of pyrimidines or position 7of7-deazapurines,not only because it then points out into the major groove of DNAand thus does not destabilize the duplex, but because in most cases,t he corresponding substituted 2'-deoxyribonucleoside triphosphates (dNTPs) are good substrates for DNAp olymerases and can be used in the polymerase-catalyzed synthesis of modified DNA. [2,3] Diverse modifications,i ncluding fluorophores, [4] redox [5] or spin labels, [6] reactive groups for conjugations, [7] and biomolecules (e.g., oligonucleotides [8] or proteins [9] ), have been introduced into the major groove through the enzymatic incorporation of modified nucleotides and applied in different fields.Modification or labelling of the minor groove has mostly been reported with 2'-and 4'-sugarmodified derivatives.[10-13] 2-Chloroadenine [14] and 2,6-diaminopurine [15] dNTPs are the only minor-groove base-modified nucleotides that have been reported as substrates for DNA polymerases,w hereas 2-arylamino-dATP derivatives were found to act as polymerase inhibitors.[16] Them inor groove sites of the nucleobases are difficult to modify since they are crucial both for Watson-Crick base pairing and for key minorgroove interactions with DNApolymerase that are important for extension of the chain.[17] On the other hand, 2-ethynylpyridone-C-nucleotide incorporated into DNA [18] formed as table base pair with adenine,a nd 2-(imidazolylalkylamino)purines in ONs also stabilized duplexes. [19] Because the possibility of minor-groove base labelling would be attractive for many prospective applications,for example,the mapping of DNA-protein interactions,w ee nvisaged that as mall substituent at position 2ofapurine may not fully disturb the key H-bonding interactions with the opposite base and the polymerase,a nd we report herein the first enzymatic synthesis of minor-groove base-modified DNA.Aseries of six 2-substituted dATP derivatives bearing Cl, NH 2 ,C H 3 ,v inyl, ethynyl and phenyl substituents (d R ATPs) was designed to study the effect of substituents of different bulkiness at position 2o fa denine on polymerase-mediated incorporation. While dCl ATP [14,20] and d NH2 ATP [15] were known, the others were prepared through triphosphorylation [21] of the corresponding 2'-deoxy-ribonucleosides (d R As...

show abstract

Sequenz‐spezifischer Einbau von Enzym‐Nukleotid‐Chimären durch DNA‐Polymerasen

Cited by 8 publications

References 37 publications

Highlights from the 53rd EUCHEM conference on stereochemistry, Bürgenstock, Switzerland, May 2018

Highlights from the 53rd EUCHEM conference on stereochemistry, Bürgenstock, Switzerland, May 2018

2‐Substituted dATP Derivatives as Building Blocks for Polymerase‐Catalyzed Synthesis of DNA Modified in the Minor Groove

2‐Substituted dATP Derivatives as Building Blocks for Polymerase‐Catalyzed Synthesis of DNA Modified in the Minor Groove

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