Synthesis of G‐N²‐(CH₂)₃‐N²‐G Trimethylene DNA Interstrand Cross‐Links

Abstract

The synthesis of G-N2-(CH2)3-N2-G trimethylene DNA interstrand cross-links (ICLs) in a 5′-CG-3′ and 5′-GC-3′ sequence from oligodeoxynucleotides containing N2-(3-aminopropyl)-2′-deoxyguanosine and 2-fluoro-O6-(trimethylsilylethyl)inosine is presented. Automated solid-phase DNA synthesis was used for unmodified bases and modified nucleotides were incorporated via their corresponding phosphoramidite reagent by a manual coupling protocol. The preparation of the phosphoramidite reagents for incorporation of N2-(3-aminopropyl)-2′-deoxyguanosine is reported. The high-purity trimethylene DNA interstrand cross-link product is obtained through a nucleophilic aromatic substitution reaction between the N2-(3-aminopropyl)-2′-deoxyguanosine and 2-fluoro-O6-(trimethylsilylethyl)inosine containing oligodeoxynucleotides.

Synthesis and Characterization of Site‐Specific O6‐Alkylguanine DNA‐Alkyl Transferase‐Oligonucleotide Crosslinks

“…2-F-dI phosphoramidite (Glen Research) 2-F-dI-containing modified DNA CPG column (see Gruppi et al, 2014) Argon gas Ghodke et al…”

“…Overall, the protocols presented here are valuable tools for the synthesis of site-specific DPCs, which can be further used to understand the complex repair mechanisms and TLS pathway. (Gruppi, Johnson Salyard, & Rizzo, 2014). A post-oligomerization approach was developed to achieve the synthesis of the N 2 -propargyl-dG-modified oligonucleotide template, as depicted in Figure 2.…”

Simple, High‐Yield Syntheses of DNA Duplexes Containing Interstrand DNA‐DNA Cross‐Links Between an N4‐Aminocytidine Residue and an Abasic Site

“…This is usually unacceptable from a preparative perspective. Alternatively, elegant syntheses of structurally-defined cross-linked duplexes have been devised (Balkrishnen et al, 1996; Carrette et al, 2013; Gao and Orgel, 1999; Gruppi et al, 2014; Haque et al, 2014; Harwood et al, 2000; Hentshel et al, 2012; Hong et al, 2006; Manoharan et al, 1999; Mukherjee et al, 2014; Nakatani et al, 2002; Nishimoto et al, 2013; Noll et al, 2001; O'Flaherty et al, 2013; Pujari et al, 2014; Schärer, 2005; Tomás-Gamasa et al, 2014; Ye et al, 2013), but many of these multi-step organic reaction sequences may not be practical in many of the biochemical and materials science laboratories with interests in cross-linked DNA. In the work described here, we sought a simple, benchtop procedure for the synthesis of DNA duplexes containing a site-specific interstrand cross-link that employs inexpensive commercially available enzymes, chemicals, and oligodeoxynucleotides.…”