peptide beyond the JmjC-domain are also observed, whereby the zinc-binding
domain of KDM6A/B, located C-terminal to JmjC-domain, provides additional
affinity and specificity [49, 51] (Fig. 6b). This region undergoes a conformational
change upon H3 peptide binding, to expose hydrophobic patches that interact with
H3(17–21) region of the peptide. These key interactions have been supported by
kinetic studies on mutations to H3 and KDM6A [49], providing evidence that
multiple factors contribute towards defining substrate binding and catalysis.
For other binding domains, such as the PHD-fingers, their spatial positioning and
distance relative to JmjC-domains can also influence the histone H3 binding
sequence specificity. PHD-fingers of KDM7 subfamily recognise the H3K4me3
mark, and in the case of KDM7B, the PHD-finger interacts with H3K4me3 in
H3K4me3K9me2 to allow demethylation at K9me2 site by JmjC-domain, while it
has negligible binding and catalysis with H3K9me2 alone [53]. The crystal structure
of KDM7B in complex with H3K4me3K9me2 reveals that the histone H3 binds
across the PHD-finger and JmjC-domain interface in an extended conformation (H3
(1–9)) and forms extensive interactions with both domains or individual domains
[53] (Fig. 6c). KDM7A, on the other hand, has a longer linker between the
PHD-finger and JmjC-domains and cannot demethylate at the K9 site of
H3K4me3K9me2, while it can demethylate at the K27 site of H3K4me3K27me2
[53]. The ancillary reader domain and linker length can thus contribute significantly
to the substrate selectivity of the KDM7s. Other JmjC-KDMs, such as the KDM4s
and KDM5s, are also allosterically regulated by their ancillary reader domains
(PHD-fingers, Tudor domains) [9, 19, 54–56]. It is anticipated that the histone
binding complex structures of multidomain KDM5s and KDM4s will provide
molecular insight into the substrate binding and cross-talk between the reader and
catalytic domains.
The histone H3 binding surfaces can also differ between the JmjC-KDMs. In
KDM2A, the H3 peptide ‘threads’ through a narrow channel that can only be
accommodated by Gly residues [42] (Fig. 6d). This steric constrain in the channel
provides selectivity towards a unique GG motif at H3K36 region (A29-PATGGV35). Methylated H3K36 then inserts into a deep cavity (‘cavity insertion’) and
locks the H3K36me2 peptide substrate into the active site pocket [42]. In contrast, in
KDM4A, the same H3 peptide sequence binds at the protein surface, and K36me3
inserts into a surface groove (‘surface groove’ recognition) [42–44]. This binding
mode can account for the ability of KDM4s to demethylate multiple histone substrates [7, 46], but KDM2A is highly specific for the H3K36me2.
In summary, the histone substrate recognition and selectivity are achieved
through a combination of unique features and mechanisms by JmjC-KDMs, including distinct methyllysine binding pockets, intermolecular side chain and backbone
interaction network, different substrate-binding channels and interplay with other
domains.
Inhibitors of JmjC-Containing Histone Demethylases
233
domain of KDM6A/B, located C-terminal to JmjC-domain, provides additional
affinity and specificity [49, 51] (Fig. 6b). This region undergoes a conformational
change upon H3 peptide binding, to expose hydrophobic patches that interact with
H3(17–21) region of the peptide. These key interactions have been supported by
kinetic studies on mutations to H3 and KDM6A [49], providing evidence that
multiple factors contribute towards defining substrate binding and catalysis.
For other binding domains, such as the PHD-fingers, their spatial positioning and
distance relative to JmjC-domains can also influence the histone H3 binding
sequence specificity. PHD-fingers of KDM7 subfamily recognise the H3K4me3
mark, and in the case of KDM7B, the PHD-finger interacts with H3K4me3 in
H3K4me3K9me2 to allow demethylation at K9me2 site by JmjC-domain, while it
has negligible binding and catalysis with H3K9me2 alone [53]. The crystal structure
of KDM7B in complex with H3K4me3K9me2 reveals that the histone H3 binds
across the PHD-finger and JmjC-domain interface in an extended conformation (H3
(1–9)) and forms extensive interactions with both domains or individual domains
[53] (Fig. 6c). KDM7A, on the other hand, has a longer linker between the
PHD-finger and JmjC-domains and cannot demethylate at the K9 site of
H3K4me3K9me2, while it can demethylate at the K27 site of H3K4me3K27me2
[53]. The ancillary reader domain and linker length can thus contribute significantly
to the substrate selectivity of the KDM7s. Other JmjC-KDMs, such as the KDM4s
and KDM5s, are also allosterically regulated by their ancillary reader domains
(PHD-fingers, Tudor domains) [9, 19, 54–56]. It is anticipated that the histone
binding complex structures of multidomain KDM5s and KDM4s will provide
molecular insight into the substrate binding and cross-talk between the reader and
catalytic domains.
The histone H3 binding surfaces can also differ between the JmjC-KDMs. In
KDM2A, the H3 peptide ‘threads’ through a narrow channel that can only be
accommodated by Gly residues [42] (Fig. 6d). This steric constrain in the channel
provides selectivity towards a unique GG motif at H3K36 region (A29-PATGGV35). Methylated H3K36 then inserts into a deep cavity (‘cavity insertion’) and
locks the H3K36me2 peptide substrate into the active site pocket [42]. In contrast, in
KDM4A, the same H3 peptide sequence binds at the protein surface, and K36me3
inserts into a surface groove (‘surface groove’ recognition) [42–44]. This binding
mode can account for the ability of KDM4s to demethylate multiple histone substrates [7, 46], but KDM2A is highly specific for the H3K36me2.
In summary, the histone substrate recognition and selectivity are achieved
through a combination of unique features and mechanisms by JmjC-KDMs, including distinct methyllysine binding pockets, intermolecular side chain and backbone
interaction network, different substrate-binding channels and interplay with other
domains.
Inhibitors of JmjC-Containing Histone Demethylases
233