Peptide nucleic acids (PNAs)
Peptide nucleic acids (PNAs) are artificially synthesized polymers which mimic DNA or RNA. Indeed, PNAs have a peptide-like backbone with nucleobases at side chains. PNAs can hybridize with both DNA and RNA sequences to form PNA/DNA and PNA/RNA structures. PNAs involve a remarkably high affinity, specificity, flexibility and stability to the target DNA or RNA. Thanks to their advantages, PNAs can be used in a variety of applications.
The PNA backbone provides an electrically neutral charge to the PNA. Consequently, the hybridization between PNA and the targeted nucleic acid sequence is facilitated due to a lack of electrostatic repulsion. So, the binding affinity is stronger and more stable for PNA/DNA than the natural homo- or heteroduplexes hybridization.
Specificity and sensitivity
PNAs show higher specificity to bind complementary DNA or RNA than natural complex like DNA/DNA. Moreover, a mismatch between PNA/DNA sequence binding is easier to identify. Indeed, the difference in binding affinity of PNA/DNA between good match and mismatch is bigger than the difference between good match and mismatch of DNA/DNA binding.
Usually, DNA sequence for hybridization have to contain 25-30 bases but thanks to PNA higher binding strength, the hybridization of PNA with DNA requires less bases (13-18 bases). The length of PNA sequence guarantee only the specificity.
Unlike DNA, PNA is quite stable under high temperature or high pH condition. Besides, the synthetic backbone of PNAs involves that PNAs are not easily recognized by nucleases, proteases or polymerase. PNAs are resistant to degradation by enzymes.
Properties and advantages of PNAs are very attractive for a variety of applications. PNA properties are useful in molecular biology procedures and diagnostic assays. Indeed, PNAs are used to detect anomaly and to develop antigens and anticancer drugs.
|Detection of genetic anomaly with PNA microarray combined with PCR – biosensors||Antigen and antisense drugs|
|Inhibition of mutant sequence with PNA clamp||Inhibition of miRNA|
|Detection of DNA or RNA sequence with fluorescent PNA – Imaging probes and FISH||Capture of DNA or RNA unwanted|
SB-PEPTIDE is able to produce peptide nucleic acids and has a custom PNAs synthesis service.
1- Pellestor F and Paulasova P. Eur J Hum Genet.12(9):694-700 (2004)
Peptide nucleic acids (PNAs) are synthetic mimics of DNA in which the deoxyribose phosphate backbone is replaced by a pseudo-peptide polymer to which the nucleobases are linked. PNAs hybridize with complementary DNAs or RNAs with remarkably high affinity and specificity, essentially because of their uncharged and flexible polyamide backbone. The unique physico-chemical properties of PNAs have led to the development of a variety of research assays, and over the last few years, the use of PNAs has proven their powerful usefulness in molecular biology procedures and diagnostic assays. The more recent applications of PNA involve their use as molecular hybridization probes. Thus, several sensitive and robust PNA-dependent methods have been designed for developing antigene and anticancer drugs, modulating PCR reactions, detecting genomic mutation or labelling chromosomes in situ.
2- Muratovska A et al. Nucleic Acids Res. 29(9):1852-63 (2001)
Targeting peptide nucleic acid (PNA) oligomers to mitochondria within cells by conjugation to lipophilic cations: implications for mitochondrial DNA replication, expression and disease
The selective manipulation of mitochondrial DNA (mtDNA) replication and expression within mammalian cells has proven difficult. One promising approach is to use peptide nucleic acid (PNA) oligomers, nucleic acid analogues that bind selectively to complementary DNA or RNA sequences inhibiting replication and translation. However, the potential of PNAs is restricted by the difficulties of delivering them to mitochondria within cells. To overcome this problem we conjugated a PNA 11mer to a lipophilic phosphonium cation. Such cations are taken up by mitochondria through the lipid bilayer driven by the membrane potential across the inner membrane. As anticipated, phosphonium-PNA (ph-PNA) conjugates of 3.4-4 kDa were imported into both isolated mitochondria and mitochondria within human cells in culture. This was confirmed by using an ion-selective electrode to measure uptake of the ph-PNA conjugates; by cell fractionation in conjunction with immunoblotting; by confocal microscopy; by immunogold-electron microscopy; and by crosslinking ph-PNA conjugates to mitochondrial matrix proteins. In all cases dissipating the mitochondrial membrane potential with an uncoupler prevented ph-PNA uptake. The ph-PNA conjugate selectively inhibited the in vitro replication of DNA containing the A8344G point mutation that causes the human mtDNA disease ‘myoclonic epilepsy and ragged red fibres’ (MERRF) but not the wild-type sequence that differs at a single nucleotide position. Therefore these modified PNA oligomers retain their selective binding to DNA and the lipophilic cation delivers them to mitochondria within cells. When MERRF cells were incubated with the ph-PNA conjugate the ratio of MERRF to wild-type mtDNA was unaffected, even though the ph-PNA content of the mitochondria was sufficient to inhibit MERRF mtDNA replication in a cell-free system. This unexpected finding suggests that nucleic acid derivatives cannot bind their complementary sequences during mtDNA replication. In summary, we have developed a new strategy for targeting PNA oligomers to mitochondria and used it to determine the effects of PNA on mutated mtDNA replication in cells. This work presents new approaches for the manipulation of mtDNA replication and expression, and will assist in the development of therapies for mtDNA diseases.
3- Egholm M et al. Nature. 365(6446):566-8 (1993)
DNA analogues are currently being intensely investigated owing to their potential as gene-targeted drugs. Furthermore, their properties and interaction with DNA and RNA could provide a better understanding of the structural features of natural DNA that determine its unique chemical, biological and genetic properties. We recently designed a DNA analogue, PNA, in which the backbone is structurally homomorphous with the deoxyribose backbone and consists of N-(2-aminoethyl)glycine units to which the nucleobases are attached. We showed that PNA oligomers containing solely thymine and cytosine can hybridize to complementary oligonucleotides, presumably by forming Watson-Crick-Hoogsteen (PNA)2-DNA triplexes, which are much more stable than the corresponding DNA-DNA duplexes, and bind to double-stranded DNA by strand displacement. We report here that PNA containing all four natural nucleobases hybridizes to complementary oligonucleotides obeying the Watson-Crick base-pairing rules, and thus is a true DNA mimic in terms of base-pair recognition.