Uracil-Directed Ligand Tethering:  An Efficient Strategy for Uracil DNA Glycosylase (UNG) Inhibitor Development

Abstract

Uracil DNA glycosylase (UNG) is an important DNA repair enzyme that recognizes and excises uracil bases in DNA using an extrahelical recognition mechanism. It is emerging as a desirable target for small molecule inhibitors given its key role in a wide range of biological processes including the generation of antibody diversity, DNA replication in a number of viruses, and the formation of DNA strand breaks during anticancer drug therapy. To accelerate the discovery of inhibitors of UNG we have developed a uracil-directed ligand tethering strategy. In this efficient approach, a uracilaldehyde ligand is tethered via alkyloxyamine linker chemistry to a diverse array of aldehyde binding elements. Thus, the mechanism of extrahelical recognition of the uracil ligand is exploited to target the UNG active site, and alkyloxyamine linker tethering is used to randomly explore peripheral binding pockets. Since no compound purification is required, this approach rapidly identified the first small molecule inhibitors of human UNG with micromolar to submicromolar binding affinities. In a surprising result, these uracil-based ligands are found not only to bind to the active site, but also to a second noncompetitive site. The weaker noncompetitive site suggests the existence of a transient binding site for uracil during the multistep extrahelical recognition mechanism. This very general inhibitor design strategy can be easily adapted to target other enzymes that recognize nucleobases, including other DNA repair enzymes that recognize other types of extrahelical DNA bases.

Charge Transfer Complexes of Some Heterocyclic Aromatic Amines with π‐Organic Acceptors in Polar Solvents

“…On the other hand, if the acceptor concentration is kept constant, the sp values should reflect the donor strength or its basicity. As expected, the results listed in Tables (1)(2)(3)(4) show that sp values follow the electron affinity order of the acceptors p-bromoaniline > p-chloroaniline which may be ascribed to salvation which would be expected to be more extensive with small ions where the charge is more concentrated compared to that with large ions.…”

“…2 Amino-and iminopyrimidine forms are in content of various biological systems and play important role in numerous biological processes. They are included in DNA, RNA, in content of vitamins of B group, antibiotics, vessel expanding medicines, correcting heart action, stimulating metabolic processes.…”

The Catalytic Power of Uracil DNA Glycosylase in the Opening of Thymine Base Pairs

“…These observations support a mechanism in which the very earliest event in recognition by UNG involves spontaneous expulsion of the base into a transient binding site that has been detected in crystallographic studies of the herpesvirus UNG bound to pTpTpT. 6,14 Once this site is transiently occupied, the base has an opportunity to partition forward along the base flipping reaction coordinate (in the case of uracil), or alternatively, to re-enter the DNA base stack (in the case of thymine). Since the lifetime of the extrahelical base in the free DNA is so short (∼100 to 800 ns, Table 1), and UNG exhibits diffusion controlled binding kinetics, it is an improbable event that UNG encounters the extrahelical base during a bimolecular collision 1 .…”

A label-free and sensitive fluorescent method for the detection of uracil-DNA glycosylase activity

“…[35][36][37] The original design in Fig. 1A underwent fluorescence enhancement in the presence of UDG, thus it displayed fluorescence quenching accordingly when UDG inhibitors were present.…”