Switchable Protecting Strategy for Solid Phase Synthesis of DNA and RNA Interacting Nucleopeptides

Abstract

Nucleopeptides are promising nucleic acid mimetics in which the peptide backbone bears nucleobases. They can recognize DNA and RNA targets modulating their biological functions. To date, the lack of an effective strategy for the synthesis of nucleopeptides prevents their evaluation for biological and biomedical applications. Herein, we describe an unprecedented approach that enables the synthesis of cationic both homo and heterosequence nucleopeptides wholly on solid support with high yield and purity. Spectroscopic studies indicate advantageous properties of the nucleopeptides in terms of binding, thermodynamic stability and sequence specific recognition. Biostability assay and laser scanning confocal microscopy analyses reveal that the nucleopeptides feature acceptable serum stability and ability to cross the cell membrane.

Investigation of the Stereochemical-Dependent DNA and RNA Binding of Arginine-Based Nucleopeptides

“…Thus, we were able to estimate the apparent Kd values of np-1 to polyA and polydA by performing CD titration experiment with increasing concentrations of np-1 in the presence of a constant concentration of nucleic acids. As depicted in Figure 6, the apparent Kd values further corroborated the results achieved by us and other groups [37,39], indicating that cationic nucleopeptides are endowed with higher affinity for RNA (Figure 6a) with the respect to DNA molecules (Figure 6b). The CD melting profiles suggested that the dissociation of np-1/polyA and np-1/polydA was a two-state process.…”

“…Finally, hydroxyl groups with R configuration were added to the 2' carbon atoms of the remaining furanose rings, and the C-and N-terminal ends of np-1 was capped with an amino and an acetyl group respectively. The initial structure of cp-1 was constructed base on the experimentally determined 2:1 stoichiometric interaction of two nucleopeptide molecules with a single 12 mer RNA oligonucleotide [37] To finally assemble the structure of cp-1 composed of a single-stranded 12 mer Adenine RNA oligonucleotide (A12 RNA ss) and two molecules of nucleopeptide np-1, the previously obtained cp-1 complex was duplicated and the RNA chains were linked covalently. The union was made by setting one copy of cp-1 on top of the other and connecting the 3' end of one RNA strands with the 5' end of the other through a newly added phosphate group.…”

“…Diederichsen et al [34] and Mihara et al [35,36,15] reported the synthesis of α and eptides with nucleobase-functionalized side chains and the use of L-α-amino-ɣ-nucleobasetyric acid to assemble RNA binding molecules and functional peptides, respectively. More ently, we have reported new synthetic strategies to obtain both homo-and hetero-sequences of ionic nucleopeptides that exhibited advantageous properties in terms of binding, specificity, cell rmeability and delivery for RNA, DNA and PNA molecules ( Figure 1) [37,38]. Although the rapeutic potential of nucleopeptides is less known with the respect to the parent PNA molecules, y are under investigation for their ability to interfere with the Reverse Transcriptase of the Human munodeficiency Virus (RT-HIV), which is fundamental for HIV infection [39].…”

Programmed Recognition between Complementary Dinucleolipids To Control the Self‐Assembly of Lipidic Amphiphiles

“…After deprotection of the amino groups in ( 3 ) with 4 m HCl in 1,4‐dioxane, which yielded the oleyl β‐aminoalanine ester derivative ( 4 ), an EDC‐based coupling reaction with thymine‐1‐acetic acid ( 5 ) resulted in the formation of dinucleolipid 1 T . For the synthesis of dinucleolipid 1 A , 6‐ N ‐ tert ‐butoxycarbonyl‐9‐carboxymethyladenine ( 6 ) was used as building block in a similar coupling reaction, and so a final step of deprotection of the adenine amino group in compound 7 with trifluoroacetic acid was necessary.…”

“…The first synthetic step consisted of an esterification of the N-Boc protected b-aminoalanine derivative(2) [34] with oleyl alcohol, mediated by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimideh ydrochloride (EDC). After deprotection of the amino groups in (3)w ith 4 m HCl in 1,4-dioxane, whichy ielded the oleyl b-aminoalanine ester derivative (4), an EDC-based couplingr eactionw ith thymine-1-acetic acid (5)r esulted in the formation of dinucleolipid 1T.F or the synthesis of dinucleolipid 1A,6 -N-tert-butoxycarbonyl-9-carboxymethyladenine (6) [35] was used as buildingb lock in as imilar couplingr eaction, and so af inal step of deprotection of the adenine amino group in compound 7 with trifluoroacetic acid was necessary.…”