Phosphate-Methylated Oligonucleotides Past, Present and Future

Abstract

At the moment, we see a great interest for application of Anti Sense Oligonucleotides (ASOs) in order to regulate the expression of genes related to certain diseases. These nucleotides obtained a number of fascinating properties by means of chemical manipulation of natural DNA and RNA under conservation of Watson-Crick base-pairing. About 35 years ago for our research in this field, we selected synthetically (short) phosphate-methylated DNA and RNA. It was concluded that there is an exclusive selection in hybridization affinity with natural DNA and RNA. These (bio)chemical and physical-chemical properties are extensively published. ASOs have found their way in public health as is clearly shown in the treatment of (progressive) neurological diseases. We focus specifically on the past, present and future of the phosphate-methylated oligonucleotides, illustrated with different research studies in chemistry and biophysics. A new field of application of modified DNAs is based on interactive improvements of sensitivity and specificity of nanowire field effect transistor gene chip by designing phosphate-methylated DNA as probe.

The Biochemical Impact by Covalent Shielding of the Anionic Oxygen of the Phosphate Group in DNA and RNA as Methylated Phosphotriester Linkage on the Inhibition of DNA Duplication and on HIV-1 RNA Viral Infectivity Has Been Seriously Overlooked

“…Recently, it was observed that DNA binding force facilitates Z-DNA formation under physiological salt conditions [2]. Generally, the impact of the presence of methyl phosphotriester (MPTE) linkages of the DNA backbone results in higher nuclease stability and the benefit of lipophilic character as has been demonstrated under biological conditions [3]. The MPTE linkages also improve the duplex stability due to the reduction of the electrostatic repulsion between the strands by eliminating the negative charge in one strand [4].…”

“…After base protection, the phosphodiester backbone was methylated as described by Miller et al [18] followed by base deprotection. In the context of a recombinant DNA study, we used the E. coli pab B gene, coding for para aminobenzoate synthetase, inserted in the M13mp18 phage selecting 2 templates for relative synthesis activity of complementary phosphatemethylated 8-and 18-mers along the template position [3].…”

“…Between 1985 and 1990, my group at Eindhoven University of Technology was involved in the HIV-1 RNA research of the preparation of MPTE antisense DNA Journal of Biophysical Chemistry with 20 nucleotides for inhibition of target regions of the virus with hairpin loop (single-stranded) structures. A highly favoured structure is TAR (trans-activation response element) with six bases in the loop which was established by X-ray [3].…”

Sensitive and specific miRNA in situ hybridization using partially methylated phosphotriester antisense DNA probes

“…1 ), was investigated and applied to the DNA sequencing based on field effect transistor (Chen et al 2013). The advantages of MPTE modifications on the DNA backbone result in higher nuclease stability and the emergence of lipophilic character (Buck 2020). Moreover, MPTE modifications improve duplex hybridization affinity due to the reduction of electrostatic repulsion (reduction of a negative charge for each MPTE linkage) between the strands of an nDNA/DNA,RNA duplex (Chou et al 2022; Huang et al 2018; Kuo et al 2020; Meade et al 2014; Miller et al 1986; Moody et al 1989).…”