To uncover the function of and interplay between the mammalian cytosine modifications 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), new techniques and advances in current technology are needed. To this end, we have developed oxidative bisulfite sequencing (oxBs-seq), which can quantitatively locate 5mC and 5hmC marks at single-base resolution in genomic DNA. In bisulfite sequencing (BS-seq), both 5mC and 5hmC are read as cytosines and thus cannot be discriminated; however, in oxBS-seq, specific oxidation of 5hmC to 5-formylcytosine (5fC) and conversion of the newly formed 5fC to uracil (under bisulfite conditions) means that 5hmC can be discriminated from 5mC. a positive readout of actual 5mC is gained from a single oxBS-seq run, and 5hmC levels are inferred by comparison with a BS-seq run. Here we describe an optimized second-generation protocol that can be completed in 2 d.
Although known for more than a century, bromine trifluoride is not a common reagent in day to day research work in organic chemistry. Its tendency to react exothermically with solvents containing Lewis base elements such as water, acetone or ethers distanced it from the mind of many. Still, under the proper conditions, it can perform quite a few selective reactions which are difficult to achieve by other reagents. We discuss in this review reactions which have been published in the last 30 years. They consist of fluorinating heteroatoms, substituting carbon‐halogen bonds with carbon‐fluorine bonds, syntheses of anesthetics, construction of the trifluoromethyl (CF3) and the difluoromethylene (CF2) groups, aromatic brominations and more.
We report here on the screening of a fragment library against a G-quadruplex element in the human c-MYC promoter. The ten fragment hits had significant concordance between a biophysical assay, in silico modelling and c-MYC expression inhibition, highlighting the feasibility of applying a fragment-based approach to the targeting of a quadruplex nucleic acid.
RNA–protein
interactions are vital throughout the HIV-1
life cycle for the successful production of infectious virus particles.
One such essential RNA–protein interaction occurs between the
full-length genomic viral RNA and the major structural protein of
the virus. The initial interaction is between the Gag polyprotein
and the viral RNA packaging signal (psi or Ψ), a highly conserved
RNA structural element within the 5′-UTR of the HIV-1 genome,
which has gained attention as a potential therapeutic target. Here,
we report the application of a target-based assay to identify small
molecules, which modulate the interaction between Gag and Ψ.
We then demonstrate that one such molecule exhibits potent inhibitory
activity in a viral replication assay. The mode of binding of the
lead molecules to the RNA target was characterized by 1H NMR spectroscopy.
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