K289M is a variant of DNA polymerase β (pol β) that has previously been identified in colorectal cancer. The expression of this variant leads to a 16-fold increase in mutation frequency at a specific site in vivo and a reduction in fidelity in vitro in a sequence context-specific manner. Previous work shows that this reduction in fidelity results from a decreased level of discrimination against incorrect nucleotide incorporation at the level of polymerization. To probe the transition state of the K289M mutator variant of pol β, single-turnover kinetic experiments were performed using β,γ-CXY dGTP analogues with a wide range of leaving group monoacid dissociation constants (pKa4), including a corresponding set of novel β,γ-CXY dCTP analogues. Surprisingly, we found that the values of the log of the catalytic rate constant (kpol) for correct insertion by K289M, in contrast to those of wild-type pol β, do not decrease with increased leaving group pKa4 for analogues with pKa4 values of <11. This suggests that one of the relative rate constants differs for the K289M reaction in comparison to that of the wild type (WT). However, a plot of log(kpol) values for incorrect insertion by K289M versus pKa4 reveals a linear correlation with a negative slope, in this respect resembling kpol values for misincorporation by the WT enzyme. We also show that some of these analogues improve the fidelity of K289M. Taken together, our data show that Lys289 critically influences the catalytic pathway of pol β.

The total synthesis of a number of representative natural products isolated from Leucetta and Clathrina sponges containing a polysubstituted 2-aminoimidazole are described. These syntheses take advantage of the site specific metallation reactions of 4,5-diiodoimidazoles resulting in the syntheses of three different classes of Leucetta derived natural products. The cytotoxicities of these natural products, along with several precursors in MCF7 cells were determined through an MTT growth assay. For comparative purposes a series of naphthimidazole-containing family members are included. Graphical Abstract

The total syntheses of two alkaloids isolated from a marine sponge of the Leucetta sp. have been accomplished in 6 and 7 steps starting from a 4,5-diiodoimidazole derivative. Grignard mediated halogen-metal exchange was used to install the benzyl side chain. C2 substitution was accomplished via lithiation followed by quenching with trisyl azide which provided isonaamine C after hydrogenation. Isonaamidine E was then prepared from isonaamine C via introduction of the hydantoin ring by reaction with an activated parabanic acid.