Activation of p300 Histone Acetyltransferase by Small Molecules Altering Enzyme Structure:  Probed by Surface-Enhanced Raman Spectroscopy

Abstract

Reversible acetylation of nucleosomal histones and nonhistone proteins play pivotal roles in the regulation of all the DNA templated phenomenon. Dysfunction of the enzymes involved in the acetylation/deacetylation leads to several diseases. Therefore, these enzymes are the targets for new generation therapeutics. Here, we report the synthesis of trifluoromethyl phenyl benzamides and their effect on histone acetyltransferase (HAT) activity of p300. One of these benzamides, CTPB (N-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-6-pentadecyl-benzamide), was discovered as a potent activator of the p300 HAT activity. We have found that pentadecyl hydrocarbon chain of CTPB is required to activate the HAT only under certain context. Furthermore, our results show that the relative position of -CF 3 and -Cl in CTB (N-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-benzamide) is also very critical for the activation. Surface-enhanced Raman spectroscopy (SERS) of p300 and the HAT activator complexes evidently suggest that the activation of HAT activity is achieved by the alteration of p300 structure. Therefore, apart from elucidating the chemical basis for small molecule mediated activation of p300, this report also describes, for the first time, Raman spectroscopic analysis of the complexes of histone-modifying enzymes and their modulators, which may be highly useful for therapeutic applications.

A Small Molecule Activator of p300/CBP Histone Acetyltransferase Promotes Survival and Neurite Growth in a Cellular Model of Parkinson’s Disease

“…Indeed, CTPB has been consistently demonstrated to induce p300/CBP HAT activity, in which histone acetylation levels were typically assessed in vitro (Balasubramanyam et al 2003;Devipriya and Kumaradhas 2013;Devipriya et al 2010;Mantelingu et al 2007) and in vivo when coupled to a CSP carrier (Selvi et al 2008). This suggests that the CTPB-induced neurotrophic effects observed in this study may be mediated by increased p300/CBP HAT activity.…”

“…CTPB is a selective and potent small molecular activator of p300/CBP that has been shown to enhance p300/CBP HAT activity in vitro (Balasubramanyam et al 2003;Devipriya and Kumaradhas 2013;Devipriya et al 2010;Mantelingu et al 2007) and in vivo when coupled to a CSP carrier (Selvi et al 2008). To determine if CTPB significantly increases p300/CBP HAT activity in this cell line, the levels of acetylated histones were first measured by immunocytochemical staining of SH-SY5Y cells for pAcH3 following CTPB treatment.…”

“…CTPB enhances p300/CBP HAT-dependent transcriptional activation, and has no effect on histone deacetylase activity (Balasubramanyam et al 2003). The ability of CTPB to selectively modulate p300 HAT activity has been further characterised (Devipriya and Kumaradhas 2013;Devipriya et al 2010;Mantelingu et al 2007), exploited to synthesize novel p300/CBP-targeting compounds (Souto et al 2010;Souto et al 2008), and demonstrated in vivo (Selvi et al 2008). However, there have been no studies investigating the phenotypic effects of CTPB-mediated p300/CBP HAT activation in cellular or animal models of PD.…”

Acetyltransferases (HATs) as Targets for Neurological Therapeutics

“…The mechanism of activation through CTPB and its derivative N-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-benzamide (CTB) was explored using surface-enhanced Raman spectroscopy, indicating a conformational change upon activator binding, which possibly recruits more acetyl-CoA and enhances auto-acetylation. Furthermore, the location of -CF3 and -Cl at the para position of the benzamide ring of CTPB and CTB is critical for HAT activation [71]. As CTPB and CTB are hydrophobic in nature, these compounds might form micelle-like structures in aqueous solution and directly bind to the hydrophobic pockets of p300.…”

Inhibition of Lysine Acetyltransferase KAT3B/p300 Activity by a Naturally Occurring Hydroxynaphthoquinone, Plumbagin

“…The amide derivative of anacardic acid, N-(4-chloro-3-trifluoromethylphenyl)-2-ethoxybenzamide), instead of being an inhibitor had HAT activation properties (30). A detailed derivatization study of these amides revealed the importance of the pentadecyl hydrocarbon chain and the presence of electronegative groups (ϪCF 3 , ϪCl) at the para position for the HAT activation (31). Curcumin, a natural p300-specific inhibitor (32), also has two hydroxyl groups.…”