Research Update
Biotin-TEG RNA Oligonucleotide Helps Dengue Virus Research
DNA Methyltransferase (MTase) methylates the C-5 position of cytosines in DNA. This is a noteworthy family of proteins due to the implications of DNA methylations in the control of gene expression. MTases are a logical target for small molecule therapeutics for cancer and other diseases. However, because the core domains of MTases are highly conserved, the elucidation of inhibitors of disease-specific MTases is a particular challenge. In a recent publication by Lim et. al. the crystal structure of dengue virus MTase with a bound S-adenosyl-L-homocysteine (SAH) derivative presented a practical path forward. SAH is the by-product of S-adenosyl-L-methionine (SAM), the MTase co-factor and methyl donor. The crystal structure revealed that the SAH N6-substituent bound in a cavity near the SAM binding site and induced conformational changes in residues lining the pocket.
This finding allowed Lim et. al. to design SAH derivatives with changes to the adenine moiety identified as responsible for inducing the conformational changes. For any considered SAH derivatives to be antiviral therapeutic candidates, Lim et. al. had to ensure that they were unreactive with human MTases. This was done using a human RNA guanine-7-MTase (hRNMT) assay, which requires a synthetic RNA substrate, a capped RNA sequence, with 3’ Biotin-TEG to stop the reaction with streptavidin coated beads when needed. This key modified RNA oligonucleotide was manufactured by TriLink and helped Lim et. al. to discover that SAH derivatives with extended phenyl rings or N-6 benzylic substituents either partially or completely lost activity against human RNA guanine-7-MTase, while effectively inhibiting the dengue virus MTase.
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Technology Spotlight
Unique Nucleic Acids Key in New High Throughput Screening Method
Dengue fever is believed to infect 50 to 100 million people worldwide annually. Rate of infection has dramatically increased by 30 fold over the past 50 years. Although there are a number of antiviral therapies on the market, there are no antidengue medications available.
In a recent publication, Niyomrattanakit et al. announced development of a high throughput screening method for elucidating effective inhibitors of a promising dengue virus drug target, RNA-dependent RNA polymerase. The team developed a fluorescence-based alkaline phosphatase–coupled polymerase assay (FAPA), which proved to be a robust tool, that could be readily adapted for other nucleotide-dependent enzymes that generate pyrophosphate. This work required modified nucleotides including a biotin labeled RNA, 3’dATP and 3’dGTP, which are offered by TriLink, in addition to many other modified nucleotides and custom synthesis services.
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Q - Can you incorporate a particular dye or modification that is not listed on your website or catalog?
A – Yes. Tell us the dye or modification you are looking to use and we will determine if it is commercially available. If it is not, there is a possibility we can make it in-house. TriLink is unique in offering both phosphoramidite synthesis and incorporation into an oligonucleotide. If neither of these is an option for you, we will research a suitable alternative modification.
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