title: Leukotriene biosynthesis inhibitor MK886 impedes DNA polymerase activity.
abstract: Specialized DNA polymerases participate in replication stress responses and in DNA repair pathways that function as barriers against cellular senescence and genomic instability. These events can be co-opted by tumor cells as a mechanism to survive chemotherapeutic and ionizing radiation treatments and as such, represent potential targets for adjuvant therapies. Previously, a high-throughput screen of ∼16,000 compounds identified several first generation proof-of-principle inhibitors of human DNA polymerase kappa (hpol κ). The indole-derived inhibitor of 5-lipoxygenase activating protein (FLAP), MK886, was one of the most potent inhibitors of hpol κ discovered in that screen. However, the specificity and mechanism of inhibition remained largely undefined. In the current study, the specificity of MK886 against human Y-family DNA polymerases and a model B-family DNA polymerase was investigated. MK886 was found to inhibit the activity of all DNA polymerases tested with similar IC(50) values, the exception being a 6- to 8-fold increase in the potency of inhibition against human DNA polymerase iota (hpol ι), a highly error-prone enzyme that uses Hoogsteen base-pairing modes during catalysis. The specificity against hpol ι was partially abrogated by inclusion of the recently annotated 25 a.a. N-terminal extension. On the basis of Michaelis-Menten kinetic analyses and DNA binding assays, the mechanism of inhibition by MK886 appears to be mixed. In silico docking studies were used to produce a series of models for MK886 binding to Y-family members. The docking results indicate that two binding pockets are conserved between Y-family polymerases, while a third pocket near the thumb domain appears to be unique to hpol ι. Overall, these results provide insight into the general mechanism of DNA polymerase inhibition by MK886.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205-7199, USA.
authors.forename: Amit
authors.initials: A
authors.lastname: Ketkar
authors.forename: Maroof K
authors.initials: MK
authors.lastname: Zafar
authors.forename: Leena
authors.initials: L
authors.lastname: Maddukuri
authors.forename: Kinrin
authors.initials: K
authors.lastname: Yamanaka
authors.forename: Surajit
authors.initials: S
authors.lastname: Banerjee
authors.forename: Martin
authors.initials: M
authors.lastname: Egli
authors.forename: Jeong-Yun
authors.initials: JY
authors.lastname: Choi
authors.forename: R Stephen
authors.initials: RS
authors.lastname: Lloyd
authors.forename: Robert L
authors.initials: RL
authors.lastname: Eoff
mesh_terms: D001665:Binding Sites; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D007700:Kinetics; D015289:Leukotrienes / Q000378:metabolism*; D016859:Lipoxygenase Inhibitors / Q000494:pharmacology*; D062105:Molecular Docking Simulation; D019384:Nucleic Acid Synthesis Inhibitors; D000094769:DNA Polymerase iota; D002722:Chlorobenzenes
chemical_list: D004259:DNA-Directed DNA Polymerase; D007211:Indoles; D015289:Leukotrienes; D016859:Lipoxygenase Inhibitors; D019384:Nucleic Acid Synthesis Inhibitors; D000094769:DNA Polymerase iota; C060893:MK-886; D002722:Chlorobenzenes; C000722070:POLI protein, human