In this review we focus on the structure and

In this review, we focus on the structure and function of the KDM4A protein, its role in cancer development, and the importance of this enzyme as a therapeutic target. For further review of the KDM4 family, see Shi et al., Whetstine et al., and Berry et al. , , . KDM4A protein structure and enzymology The KDM4A gene is a member of the Jumonji domain 2 (JMJD2) family and encodes a protein that contains JmjC and JmjN domains that form a composite active site, two PHD-type zinc finger domains, and two hybrid Tudor domains that form a bilobal structure, with each lobe resembling a normal Tudor domain (Figure 1A) 24, 25. The function of the PHD fingers of KDM4A is not yet clear (22), in Kinase Inhibitor Library to the functions of the PHD fingers present in other proteins, such as those in the NURF complex, which are known to bind to the H3K4me3 histone mark (26). The hybrid Tudor domains are formed by the exchange of the β3 and β4 chains; therefore, the electrostatic potential of the second Tudor domain is more negative than that of the first domain 21, 27, 28. Because of the folding of the hybrid Tudor domains of KDM4A, the side chain of H3K4me3 is inserted into the aromatic cage pocket of one Tudor domain, whereas the side chains of the other Tudor domain form intermolecular contacts; these domains also bind H4K20me3 peptides but in the opposite direction 28, 29, 30, 31, 32. Additionally, in vitro assays have demonstrated that this enzyme can demethylate di- and trimethylated residues at lysines 9 and 36 of histone 3 (H3K9me3/2 and H3K36me3/2, respectively), but this enzyme cannot demethylate monomethylated residues; in vivo however, KDM4A demethylates only trimethylated residues (18). KDM4A also has a higher affinity for H3K9me3 than for H3K36me3 21, 27, 28. In particular, the H3K9me3 mark is associated with heterochromatic regions and transcriptional repression (8). Although H3K36me3 is associated with transcriptional repression in some models, it is primarily involved in transcription elongation by the RNA polymerase II, transcription initiation, alternative splicing, and DNA repair and recombination. For further review, see Pradeepa et al. (33). Interestingly, the unusual KDM4A specificity for two regions with different sequences can be explained because the interplay between the enzyme and the histone peptides is governed by weak interactions such as hydrogen bonds and van der Waals interactions and by interactions with substrate backbone peptides 18, 34. In addition, the N-terminal residues of H3K9me3 and H3K36me3 share a similar β-chain conformation, and the peptides bind in the same direction within the substrate-binding site 18, 34. Thus, the trimethyl lysine is deposited in the catalytic site, which has an Fe(II) ion that is essential for the catalytic activity of the enzyme 18, 34. The proposed reaction mechanism of KDM4A is very similar to that of other Fe(II)-containing and α-ketoglutarate–dependent hydroxylases (Figure 1B). This process involves five general steps (35). (1) First, the active unbound Fe(II) ion is in a +2 oxidation state and is coordinated by two histidine residues, one glutamate residue, and three molecules of water. (2) Second, the α-ketoglutarate and diatomic oxygen are coordinated to the iron center, displacing the water molecules. (3) Third, a single electron transfer occurs from the Fe(II) ion to the oxygen molecule, leading to the formation of a peroxide radical that attacks the α-ketoglutarate and yields a mixed anhydride that is attached to the Fe3+–hydroxyl radical. (4) Fourth, this highly reactive radical activates the carbon–hydrogen bond of the methyl group located on the methyl lysine by removing a proton and transferring the hydroxyl group to the carbon atom of the methyl group, leading to hydroxymethyl lysine formation. (5) Finally, the demethylation reaction proceeds with the spontaneous loss of formaldehyde from the hydroxymethyl lysine because the carbonyl is a good leaving group. Due to the hydroxyl group transfer, which leaves a gap in the coordination sphere of the Fe2+, the mixed anhydride dissociates, producing succinate and carbon dioxide as byproducts. The union of three water molecules with the Fe2+ regenerates the original catalytic site (35).