Spacers refer to a kind of linkers that play a role of space separation when peptide nucleic acid (PNA) couple peptides or dyes.
PNA is a synthetic polymer similar to DNA or RNA. Similar to polypeptides, various purine and pyrimidine bases are connected to the backbone by methylene carbonyl groups. Because PNA is not negatively charged and has no electrostatic repulsion with DNA and RNA, the stability and specificity of PNA binding are greatly improved. It is not easy to be hydrolyzed by DNase and protease, and PNA is extremely stable in vivo and in vitro.
Spacers are designed to add space either within an oligo sequence or between the oligo and a label. Therefore, spacers can be used to separate space when PNA couple peptides or dyes. Meanwhile, multiple incorporations of varying lengths of spacers allow the precise length of the spacer arm to be controlled. This can be important in the coupling of PNA.
The main chain of PNA is linked together based on polyamide, and PNA can easily combine with polypeptide to increase function. For example, the addition of lysine can improve the solubility of PNA, and the addition of cysteine allows PNA to be linked with other molecules by disulfide bonds. Both ends of the PNA chain can be used to couple peptides, but they are often coupled at the 5' end (peptide+PNA). Spacers, which play the role of space separation, shall be added between polypeptide and PNA.
Spacers can offer a variety of spacer phosphoramidites. These are often used to distance a modifier from the oligonucleotide sequence to reduce the possibility of any adverse interaction between the modifier and the sequence. For instance, G-rich sequences are known to quench fluorescein, therefore the use of a suitable spacer will remove the dye label from the proximity of the oligonucleotide minimizing the quenching effect. In a similar fashion, spacers can be used to add distance between PNA and self-quenching dyes.