Synthesis and nucleic acid binding properties of novel peptide nucleic acids carrying beta amino acid spacer

Abstract

A synthetic method for novel Peptide Nucleic Acid (PNA) carrying Beta-amino acid spacers has been developed. The PNA monomers in this research were pyrrolidine derivatives modified with 4 natural nucleobases at C-4 position. The thymine monomers were synthesized in all possible stereoisomers including "cis-D", "trans-D", "cis-L" and "trans-L". The adenine, cytosine and guanine monomers were synthesized only in "cis-D" configuration. These PNA monomers were coupled with Beta-amino acid spacers in an alternate fashion by Fmoc solid phase peptide synthesis. The spacers included in this studies were classified into three categories: acyclic (amino-oxy acetic acid, N-amino N-methylglycine), cyclic with one stereogenic center (D-aminopyrrolidine carboxylic acid or D-APC, L-aminopyrrolidine carboxylic acid or (L-APC), cyclic with two stereogenic centers [(1S,2S), (1R,2R), (1S,2R) and (1R,2S)-2-aminocylopentane carboxylic acids (ACPC)]. The solid-phase synthesis proceeded efficiently with more than 90% average efficiency for each individual coupling step. PNA sequences in 5-10 base lengths were successfully synthesized and were characterized by MALDI-TOF MS. Binding properties of these PNA with DNA were studied by UV-titration, T[subscript m], gel electrophoresis, circular dichroism and fluorescence experiments. These revealed that the cis-D pyrrolidine PNA containing the D-aminopyrrolidine carboxylic acid (D-APC) or (1S,2S)-2-aminocyclopentane carboxylic acid [(1S,2S)-ACPC] spacers can form stable 1:1 hybrids with DNA. The PNA-DNA interactions followed the Watson-Crick base-pairing rules (A.T, C.G). The cis-D/D-APC PNA also exhibited a strong preference for binding to DNA in antiparallel fashion.