Glycol nucleic acids as duplex scaffold for the design of self-assembeld and self-organized architectures

In this thesis, GNA is explored as a simplified duplex scaffold for arranging different chromophores and the properties of the resulting chromophore assemblies are investigated. Chromophore nucleotides were incorporated into GNA by automated solid phase synthesis of oligonucleotides. The synthes...

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Bibliographic Details
Main Author: Zhou, Hui
Contributors: Zhang, Lilu (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2011
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Summary:In this thesis, GNA is explored as a simplified duplex scaffold for arranging different chromophores and the properties of the resulting chromophore assemblies are investigated. Chromophore nucleotides were incorporated into GNA by automated solid phase synthesis of oligonucleotides. The synthesis of different chromophore glycol nucleoside GNA building blocks was presented in chapter 2. The key step for synthesizing these glycol nucleosides was the regioselective and stereospecific ring-opening of dimethoxytrityliated (S)-glycidol (3). As shown in Scheme 7.1A, compounds 5, 9, 14 and 17 were synthesized by three efficient methods, namely, the application of Grignard reagents, Grignard reagents containing a metallation/transmetallation protocol, and organolithum reagents. Among these compounds, compound 17c can be also synthesized by Pd-catalyzed Sonogashira coupling reaction of compound 30 with compound 17d (Scheme 7.1 B), thus any chromophore with alkyne groups can be introduced to GNA by this method. In chapter 3, we investigated GNA duplexes containing fluorescent pyrene (Pyr) and pyrene acetylide nucleotides (Pyr′) (Figure 7.1A). Duplexes with one or two adjacent Pyr:Me or Pyr′:H base pairs were synthesized, all resulting in thermally stable duplexes at room temperature. The incorporation of pyrene or pyrene acetylide to GNA duplexes did not distort the overall GNA duplex structure. Interestingly, only the pyrene acetylides but not the related pyrene nucleotides, could form strong excimers upon interstrand stacking within the GNA duplexes (Figure 7.1B). The reason for this may be a combination of structural effects, influence of the stacking of adjacent pyrene acetylides nucleobases, and electronic effects due to the conjugation of the aromatic pyrene with the acetylide π-system. As an application of excimer emission upon duplex formation with pyrene acetylide containing GNA strands, we developed a metal ion sensor by additionally incorporating a metal-mediated base pair: hydroxypyridone homo-base pair (M:M) or hydroxypyridone-pyridylpurine hetero-base pair (M:P). As shown in Figure 7.2, D15 and D19 are very sensitive and selective Cu2+ “turn-on” fluorescent sensors. Unfortunately, our attempt to develop a Cu2+ sensor for a complex biological environment failed. In chapter 4, the porphyrin acetylide nucleotide (P) was incorporated into GNA duplexes opposite ethylene glycol abasic sites and the duplexes were analyzed by UV-melting, UV-vis, fluorescence spectroscopy, and circular dichroism. The modified duplexes display lower thermal stabilities, however the thermal stabilities of duplex containing P:H base pairs could be modulated by the incorporation of zinc(II), manganes(II) or nickel(II) ions into P (Figure 7.3B). The incorporation of zinc(II) or manganes(II) ion led to a decrease in duplex stability, but the incorporation of nickel(II) resulted in increased duplex stability. These modulations of duplex stabilities by the nature of the metal ion can be interpreted by their differenct coordination behavior. Zn2+ or Mn2+ prefers to coordinate to axial ligands when incorporated into a porphyrin in an octahedral fashion and needs to dissociate these ligands if it wants to stack within GNA between neighboring base pairs. In contrast, Ni2+ prefers square planar coordination and can therefore be accommodated easily in the base stacking. Furthermore, GNA duplexes provided a suitable scaffold to bring two porphyrins into close contact and to allow the interaction of porphyrins with different coordinated metal ions. The obvious change of the Soret bands accompanied by a blue shift (Figure 7.4B), together with the decrease of fluorescence intensity upon duplex formation revealed that a ground state interaction between two porphyrin moieties occurred in GNA, performing in face-to-face (H-dimer) fashion. In chapter 5, Electron donor-acceptor chromophores systems, mainly composed of perylene bisimide (PBI) and porphyrin (P) units were organized in duplex GNA (Figure 7.5A). The GNA duplexes which contain B-Pyr, B-Pyr′, B-Pe′ and B-P were investigated. The thermal stabilities revealed that B-P pair stabilizes the GNA duplex the most significantly, exceeding the stability of the native GNA duplex by additional 10 oC. The significantly increased duplex stability together with the red-shifted absorption band and induced CD signal at Soret band indicated the interstrand π-π stacking of B-P pair in the duplex. Subsequently, we investigated the systems which contain a couple of porphyrin and PBI building blocks. As shown in Figure 7.6B, the intensity of the CD signal accompanying the blue-shift of the bisignate CD spectra in the Soret band increased with the increasing amount of chromophore moieties. This continuous shift in a CD spectrum with increased amounts of chromophores indicated the interaction between PBI and porphyrin resulted in a highly ordered helical structure. In addition, we also synthesized some single strands which consisted of entirely PBIs or porphyrins. However, combining these single strands did not give us desired helical structure according to CD analysis. It seems that flanking natural base pairs or natural overhangs play a key role in affording helical structures. Considering the significantly decreased yield of modified GNA strand with multi-incorporated chromophores, photochemical ligation of GNA was developed in order to address this problem. In chapter 6, the photochemical ligation of GNA via anthracene cyclodimer formation was explored. Anthrancenes were introduced into GNA not only as artificial nucleosidic bases but also as phototriggered joints (Figure 7.7A). Upon the duplex formation, the two adjacent anthracene nucleobases get close to each other, and the duplex was expected to generate a hairpin structure via anthracene cyclodimer formation as a result of the photoirradiation (366 nm). The photochemical ligation of GNA via anthracene cyclodimer formation was investigated in terms of three aspects, namely, template-supported GNA ligation, interstrand crosslinking in the middle of the GNA duplex, and end capping of the GNA duplex. The template-supported GNA photoligation did not occur, probably due to the inefficient dimerization of anthracenes in tandem duplex. However, both the crosslinking in the middle of duplex and end capping of duplex proved to be feasible. Resulting intramolecular duplexes exhibited high melting temperature (Figure 7.7B), which was consistent with the formation of an intramolecular duplex with a hairpin structure. In addition, the CD spectra suggested that end capping did not disrupt overall GNA duplex structure. In this study, the GNA duplex was employed as scaffold for the design of self-assembled and self-organized architectures. Several chromophores had been successfully introduced into the interior of GNA duplexes. GNA as a simplified general duplex scaffold proves to be useful for the defined chromophores organization. Future work will be focused on applications of porphyrin-GNA conjugates as well as modified GNA containing porphyrin and PBI for light harvesting and charge transport. In addition, the photochemical ligation of GNA via anthracene cyclodimer formation is a useful tool for the interstrand crosslinking of GNA, which may provide a new method for the construction of functional GNA-supramolecules.