Cholesterol hemisuccinate: A selective inhibitor of family X DNA polymerases

Cholesterol hemisuccinate (compound 5), which consists of succinic acid esterified to the β-hydroxyl group of cholesterol, selectively and strongly inhibited the activities of mammalian DNA polymerases (pols) such as pol β, pol λ, and terminal deoxynucleotidyltransferase (TdT), which are family X pols, in vitro, and the IC50 values were 2.9, 6.3, and 6.5 μM, respectively. The compound moderately suppressed the activities of other mammalian pols such as pol A (i.e., pol γ), pol B (i.e., pols α, δ, and ε), and pol Y (i.e., pols ι, η, and κ) with 50% inhibition observed at concentrations of 131, 89.2-98.0, and 120-125 μM, respectively. The compound had no influence on the activities of plant pols α and β, prokaryotic pols and other DNA metabolic enzymes tested. Since other cholesterol-related compounds such as cholesterol, cholesteryl chloride, cholesteryl bromide, cholesteryl acetate, and cholesteryl-5α, 6α-epoxide (compounds 1-4 and 6, respectively) did not influence the activities of any enzymes tested, the hemisuccinate group of compound 5 could be important for inhibition of the pol X family. Surface plasmon resonance analysis demonstrated that compound 5 bound selectively to the C-terminal 31 kDa domain of pol β and pol λ containing a pol β-like region. On the basis of these results, the inhibitory mechanism of compound 5 on the pol X family was discussed.