'Ain't it a Ripping Night': Alcoholism and the Legacies of Empire in Salman Rushdie's Midnight's Children.

In the era of decolonisation that followed the Second World War, various authors sought to engage with India and the Empire’s past anew throughout their novels, identifying medicine and illness as key parts of Imperial authority and colonial experience. Salman Rushdie’s approach to the Raj in Midnight’s Children (1981) focused on the broad sweep of colonial life, juxtaposing the political and the personal. This article argues that Rushdie explores the history of colonial India by employing alcohol and alcoholism as lenses through which to explore the cultural, political and medical legacies of Empire. Through analysis of Midnight’s Children as well as a range of medical sources related to alcohol and inebriation, it will illustrate how drinking is central to Rushdie’s approach to secular and religious identities in newly independent India, as well as a means of satirising and undermining the supposed benefit that Empire presented to India and Indians

Structural context of disease-associated mutations and putative mechanism of autoinhibition revealed by X-ray crystallographic analysis of the EZH2-SET domain.

The enhancer-of-zeste homolog 2 (EZH2) gene product is an 87 kDa polycomb group (PcG) protein containing a C-terminal methyltransferase SET domain. EZH2, along with binding partners, i.e., EED and SUZ12, upon which it is dependent for activity forms the core of the polycomb repressive complex 2 (PRC2). PRC2 regulates gene silencing by catalyzing the methylation of histone H3 at lysine 27. Both overexpression and mutation of EZH2 are associated with the incidence and aggressiveness of various cancers. The novel crystal structure of the SET domain was determined in order to understand disease-associated EZH2 mutations and derive an explanation for its inactivity independent of complex formation. The 2.00 Å crystal structure reveals that, in its uncomplexed form, the EZH2 C-terminus folds back into the active site blocking engagement with substrate. Furthermore, the S-adenosyl-L-methionine (SAM) binding pocket observed in the crystal structure of homologous SET domains is notably absent. This suggests that a conformational change in the EZH2 SET domain, dependent upon complex formation, must take place for cofactor and substrate binding activities to be recapitulated. In addition, the data provide a structural context for clinically significant mutations found in the EZH2 SET domain

EZH2-SET mutations that may affect cofactor binding.

<p>The crystal structure of the EZH2-SET domain is represented as a ribbon model (cyan) with the hypothetical positions of cofactor and substrate (sticks colored by atom: carbon, yellow; oxygen, red; nitrogen, blue) extracted from the superimposed structure of EHMT1/PEPTIDE/SAH (PDB ID: 3HNA). EZH2-SET amino acid side chains are represented as sticks colored by atom (C, cyan; O, red; N, nitrogen). Secondary structure elements are labeled. Mutations at positions A692 {(DLBCL) (A>V) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B23" target="_blank">23</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B33" target="_blank">33</a>]}, N693 {(AMML) (N>T) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B18" target="_blank">18</a>]; (ETP ALL) (N>Y) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B26" target="_blank">26</a>]; (MF) (N>Y) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B31" target="_blank">31</a>]}, and H694 {(WS) (H>Y) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B22" target="_blank">22</a>]; (CMML) (H>R) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B18" target="_blank">18</a>]} have been found in association with numerous diseases. All three mutations likely affect cofactor binding. An S695L mutation was identified in both WS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B21" target="_blank">21</a>] and ETP ALL [26]. This mutation may affect cofactor and substrate binding indirectly by influencing the conformation of residues in direct contact with these ligands. </p

Additional disease-associated mutations outside this active site.

<p>The crystal structure of the EZH2-SET domain is represented as a ribbon model (cyan) with the hypothetical positions of cofactor and substrate (sticks colored by atom: carbon, yellow; oxygen, red; nitrogen, blue) extracted from the superimposed structure of EHMT1/PEPTIDE/SAH (PDB ID: 3HNA). EZH2-SET amino acid side chains are represented as sticks colored by atom (carbon, cyan; oxygen, red; nitrogen, blue). Secondary structure elements are labeled. (<b>a</b>) A V626M mutation was identified in WS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B21" target="_blank">21</a>]. This residue is located in the loop connecting β-1 and β-2 and may indirectly affect cofactor binding. (<b>b</b>) A K639E mutation was identified in WS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B21" target="_blank">21</a>]. This residue is located in the loop connecting β-2 and β-3. (<b>c</b>) R684 mutations were identified in WS (R>C) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B21" target="_blank">21</a>], ETP ALL (R>H) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B26" target="_blank">26</a>], and MF (R>C) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B31" target="_blank">31</a>]. This residue does not participate in cofactor or substrate binding; however, its side chain does pack against α-4 which does participate in substrate binding in homologous SET domains. R690 mutations were identified in CMML (R>H) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B17" target="_blank">17</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B18" target="_blank">18</a>] and MDS (R>C) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B28" target="_blank">28</a>]. This residue packs against F670 which in homologous SET domains contributes to the substrate lysine binding channel.</p

The 2.5 Å Crystal Structure of the SIRT1 Catalytic Domain Bound to Nicotinamide Adenine Dinucleotide (NAD⁺) and an Indole (EX527 Analogue) Reveals a Novel Mechanism of Histone Deacetylase Inhibition

The sirtuin SIRT1 is a NAD⁺-dependent histone deacetylase, a Sir2 family member, and one of seven human sirtuins. Sirtuins are conserved from archaea to mammals and regulate transcription, genome stability, longevity, and metabolism. SIRT1 regulates transcription via deacetylation of transcription factors such as PPARγ, NFκB, and the tumor suppressor protein p53. EX527 (<b>27</b>) is a nanomolar SIRT1 inhibitor and a micromolar SIRT2 inhibitor. To elucidate the mechanism of SIRT inhibition by <b>27</b>, we determined the 2.5 Å crystal structure of the SIRT1 catalytic domain (residues 241–516) bound to NAD⁺ and the <b>27</b> analogue compound <b>35</b>. <b>35</b> binds deep in the catalytic cleft, displacing the NAD⁺ nicotinamide and forcing the cofactor into an extended conformation. The extended NAD⁺ conformation sterically prevents substrate binding. The SIRT1/NAD⁺/<b>35</b> crystal structure defines a novel mechanism of histone deacetylase inhibition and provides a basis for understanding, and rationally improving, inhibition of this therapeutically important target by drug-like molecules

The EZH2-SET domain C-terminus partially occupies the substrate binding groove.

<p>(a) The EZH2-SET (cyan) and hEHMT1-SET (orange) (PDB ID:3HNA) domains are superimposed and represented by ribbons. Zinc bound by hEHMT-SET is represented as a gray sphere. The substrate peptide bound by hEHMT1 is a yellow ribbon with the lysine side chain represented as sticks. The SAH bound by hEHMT1-SET is represented by sticks and colored by atom (carbon, yellow; oxygen, red; nitrogen, blue; sulfur, sienna). The C-terminal tail of EZH2-SET turns upwards and occupies the upper region of the substrate binding groove (red arrow pointing up). The C-terminus of hEHMT1-SET turns downward (red arrow pointing downward) forming the lower lobe of the cofactor binding pocket and coordinating one zinc atom. (b) The EZH2-SET (cyan) and SUV39H2 SET domain (magenta) (PDB ID:2R3A) crystal structures are superimposed and represented by ribbons. The C-termini in both structures occupy the collapsed substrate binding groove.</p

Structural context of Y646 and A682 mutations.

<p>The crystal structure of the EZH2-SET domain is represented as a ribbon model (cyan). Side chains are represented as sticks colored by atom (carbon, cyan; oxygen, red). Secondary structure elements are labeled. Y646 is completely buried in a hydrophobic cluster except for the solvent exposed tip of the phenyl ring where the phenyl oxygen forms a hydrogen bond with a water molecule. A682 is packed against the Y646 side chain distal to the catalytic site. Mutation of A682 likely indirectly affects substrate specificity by influencing the conformation of Y646 in the active state. Y646 and A682 mutations have been found in lymphoma [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B24" target="_blank">24</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B27" target="_blank">27</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B33" target="_blank">33</a>], WS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B21" target="_blank">21</a>], and AML [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B29" target="_blank">29</a>].</p

Location of mutation in the first zinc binding domain of EZH2-SET.

<p>EZH2-SET (cyan) is represented as a ribbon diagram with zinc atoms shown as gray spheres and side chain represented as sticks (carbon, cyan; nitrogen, blue; sulfur, sienna) A H530N mutation was identified in AML [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B29" target="_blank">29</a>]. This mutation disrupts coordination of zinc in the first zinc binding domain likely having a strong destabilizing effect on the protein.</p

Mutations in the β-5/β-6 loop of EZH2-SET are contiguous with the putative substrate binding cleft.

<p>The crystal structure of the EZH2-SET domain is represented as a ribbon model (cyan). Side chains are represented as sticks colored by atom (carbon, cyan; oxygen, red; nitrogen, blue). Secondary structure elements are labeled. N673, L674, and N675 all interact directly with the C-terminal tail which occupies the substrate binding groove. Mutation of these residues could potentially affect substrate binding in the active state as well as the transition from the inactive to active state. An N673S mutation has been identified in CMML [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B32" target="_blank">32</a>]. L674V mutations have been found in both MDS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B28" target="_blank">28</a>] and AML [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B29" target="_blank">29</a>]. An N675K mutation was discovered in RCMD [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B28" target="_blank">28</a>].</p

Location of mutation in the second zinc binding domain of EZH2-SET.

<p>EZH2-SET (cyan) is represented as a ribbon diagram with zinc atoms shown as gray spheres and side chain represented as sticks (carbon, cyan; nitrogen, blue; sulfur, sienna) A C571Y mutation was identified in MF [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B31" target="_blank">31</a>] and a C576W mutation was identified in MDS [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B20" target="_blank">20</a>]. These mutations disrupt coordination of zinc in the second zinc binding domain likely destabilizing the protein. Additionally, a P577L mutation was observed in ETP ALL [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0084147#B26" target="_blank">26</a>].</p