Photoligation of self-assembled DNA constructs containing anthracene-functionalized 2 0 -amino-LNA monomers Karol Pasternak a,  , Anna Pasternak a,  , Pankaj Gupta a,  , Rakesh N. Veedu a,b,  , Jesper Wengel a,⇑ a Nucleic Acid Center, Department of Physics and Chemistry, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark b School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, Brisbane, Queensland 4072, Australia article info Article history: Received 7 July 2011 Revised 13 October 2011 Accepted 17 October 2011 Available online 20 October 2011 Keywords: Amino-LNA Anthracene Photoligation DNA cross-linking abstract Efficient synthesis of a novel anthracene-functionalized 2 0 -amino-LNA phosphoramidite derivative is described together with its incorporation into oligodeoxynucleotides. Two DNA strands with the novel 2 0 -N-anthracenylmethyl-2 0 -amino-LNA monomers can be effectively cross-linked by photoligation at 366 nm in various types of DNA constructs. Successful application of three differently functionalized 2 0 -amino-LNA monomers in self-assembled higher ordered structures for simultaneous cross-linking and monitoring of assembly formation is furthermore demonstrated. Ó 2011 Elsevier Ltd. All rights reserved. 1. Introduction Chemical ligation and photodimerization reactions have been implemented within synthetic chemistry, biochemistry, biomedi- cine, and nanotechnology. 1–10 Accordingly, there are numerous re- ports concerning both chemical ligation 3,7,8,11–14 and photoligation of nucleic acid constructs. 15–21 Photoligation can be considered the more promising of these two non-enzymatic reactions for covalent linking of nucleic acid strands as it does not require any reagents except for the nucleic acid strands involved. Moreover, the reaction progress can be straightforwardly controlled by the duration of sample irradiation and the wavelength applied. 3,15 As nucleic acids can be damaged by UV-B light (280–315 nm) 22 it is important for potential in vivo applications to identify suitable photo-reactive groups which are able to form photoadducts at a wavelength safe for human cells. One such group of compounds that are photoac- tive above 300 nm are 9-substituted anthracene derivatives. Anthracene has been applied as an intercalator and fluorophore in nucleic acid contexts 23,24 and is known to form photodimers after exposure to light of a wavelength of 366 nm. 15 There are only few reports describing formation of covalent bonds between nucleic acid strands via anthracene photo-dimer forma- tion, 15,16,18 and photoligation between anthracene rings has mostly involved derivatives attached via linkers to oligonucleotide 5 0 - and 3 0 -ends. We decided to combine the photochemical properties of 9- substituted anthracene derivatives with the duplex-stabilizing and conjugating properties of 2 0 -amino-LNA monomers 25,26 to introduce covalent junctions either in the middle or at terminal positions of complementary oligonucleotides. Notably, the 2 0 -amino group of 2 0 -amino-LNA monomers gives the opportunity of introducing addi- tional functional groups which has resulted in numerous novel oligo- nucleotide derivatives with various interesting properties and applications. 26–32 Herein we report efficient synthesis of a novel 2 0 - amino-LNA monomer functionalized with an anthracen-9-ylmethyl group and the versatility of such monomers for photochemically in- duced cross-linking of complementary nucleic acid strands. 2. Results and discussion 2.1. Chemical synthesis of anthracene-functionalized 2 0 -amino- LNA phosphoramidite 3 5 0 -O-(4,4 0 -Dimethoxytrityl)-protected 2 0 -amino-LNA nucleoside 1 was prepared according to previously described procedures. 33 Reductive amination of derivative 1 with 9-anthraldehyde in the presence of sodium triacetoxyborohydride 34 gave nucleoside 2 in 76% yield. Subsequent standard phosphitylation using 2-cyano- ethyl-N,N,N 0 ,N 0 -tetraiso-propylphosphordiamidite furnished in 69% yield the anthracene-functionalized 2 0 -amino-LNA phospho- ramidite 3 (Scheme 1) which was used on an automated DNA syn- thesizer to prepare oligonucleotides containing monomer T / (Table 1; Scheme 2; See Section 4 for details). 2.2. Photoligation studies The sequences of the oligonucleotide substrates used in photo- ligation reactions and the expected constructs formed upon 0968-0896/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmc.2011.10.052 ⇑ Corresponding author. Tel: +45 6550 2510; fax: +45 6550 4385. E-mail address: jwe@ifk.sdu.dk (J. Wengel).   Tel.: +45 6550 2510; fax: +45 6617 8760. Bioorganic & Medicinal Chemistry 19 (2011) 7407–7415 Contents lists available at SciVerse ScienceDirect Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc