Enzyme kinetics and inhibition of histone acetyltransferase KAT8.
Lysine acetyltransferase 8 (KAT8) is a histone acetyltransferase (HAT) responsible for acetylating lysine 16 on histone H4 (H4K16) and plays a role in cell cycle progression as well as acetylation of the tumor suppressor protein p53. Further studies on its biological function and drug discovery initiatives will benefit from the development of small molecule inhibitors for this enzyme. As a first step towards this aim we investigated the enzyme kinetics of this bi-substrate enzyme. The kinetic experiments indicate a ping-pong mechanism in which the enzyme binds Ac-CoA first, followed by binding of the histone substrate. This mechanism is supported by affinity measurements of both substrates using isothermal titration calorimetry (ITC). Using this information, the KAT8 inhibition of a focused compound collection around the non-selective HAT inhibitor anacardic acid has been investigated. Kinetic studies with anacardic acid were performed, based on which a model for the catalytic activity of KAT8 and the inhibitory action of anacardic acid (AA) was proposed. This enabled the calculation of the inhibition constant Ki of anacardic acid derivatives using an adaptation of the Cheng-Prusoff equation. The results described in this study give insight into the catalytic mechanism of KAT8 and present the first well-characterized small-molecule inhibitors for this HAT.
Department of Pharmaceutical Gene Modulation, Groningen Research Institute of Pharmacy, University of Groningen, Groningen, The Netherlands.
Hannah
H
Wapenaar
Department of Pharmaceutical Gene Modulation, Groningen Research Institute of Pharmacy, University of Groningen, Groningen, The Netherlands.
Petra E
PE
van der Wouden
Department of Drug Design, Groningen Research Institute of Pharmacy, University of Groningen, Groningen, The Netherlands.
Matthew R
MR
Groves
Department of Drug Chemistry and Technologies, 'Sapienza' University, Rome, Italy.
Dante
D
Rotili
Department of Drug Chemistry and Technologies, 'Sapienza' University, Rome, Italy; Pasteur Institute, Cenci Bolognetti Foundation, 'Sapienza' University, Rome, Italy.
Antonello
A
Mai
Department of Pharmaceutical Gene Modulation, Groningen Research Institute of Pharmacy, University of Groningen, Groningen, The Netherlands. Electronic address: f.j.dekker@rug.nl.
Frank J
FJ
Dekker
D000105:Acetyl Coenzyme A / Q000378:metabolism; D048729:Anacardic Acids / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D055162:Biocatalysis / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D051548:Histone Acetyltransferases / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D013329:Structure-Activity Relationship
D048729:Anacardic Acids; D004791:Enzyme Inhibitors; D006657:Histones; C088115:anacardic acid; D000105:Acetyl Coenzyme A; D051548:Histone Acetyltransferases; C496552:KAT8 protein, human
Catalytic mechanism; Enzyme kinetics; Epigenetics; Histone acetylation; KAT8