Lipid-linked nucleoside triphosphates for enzymatic synthesis of hydrophobic oligonucleotides with enhanced membrane anchoring efficiency
Abstrakt
We designed and synthesized a series of 2 '-deoxyribonucleoside triphosphates (dNTPs) bearing various lipid moieties. Fatty acid- and cholesterol-modified dNTPs proved to be substrates for KOD XL DNA polymerase in primer extension reactions.
They were also mutually compatible for simultaneous multiple incorporations into the DNA strand. The methodology of enzymatic synthesis opened a pathway to diverse structurally unique lipid-ON probes containing one or more lipid units.
We studied interactions of such probes with the plasma membranes of live cells. Employing a rational design, we found a series of lipid-ONs with enhanced membrane anchoring efficiency.
The in-membrane stability of multiply modified ONs was superior to that of commonly studied ON analogues, in which a single cholesterol molecule is typically tethered to the thread end. Notably, some of the probes were detected at the cell surface even after 24 h upon removal of the probe solution.
Such an effect was general to several studied cell lines.