Influence of the disordered domain structure of MeCP2 on its structural stability and dsDNA interaction

Abstract

Methyl-CpG binding protein 2 (MeCP2) is a transcriptional regulator and a chromatin-associated structural protein. MeCP2 deregulation results in two neurodevelopmental disorders: MeCP2 dysfunction is associated with Rett syndrome, while excess of activity is associated with MeCP2 duplication syndrome. MeCP2 is an intrinsically disordered protein (IDP) constituted by six structural domains with variable, small percentage of well-defined secondary structure. Two domains, methyl-CpG binding domain (MBD) and transcription repressor domain (TRD), are the elements responsible for dsDNA binding ability and recruitment of the gene transcription/silencing machinery, respectively. Previously we studied the influence of the completely disordered, MBD-flanking domains (N-terminal domain, NTD, and intervening domain, ID) on the structural and functional features of the MBD (Claveria- Gimeno, R. et al. Sci Rep. 2017, 7, 41,635). Here we report the biophysical study of the influence of the remaining domains (transcriptional repressor domain, TRD, and C-terminal domains, CTDα and CTDβ) on the structural stability of MBD and the dsDNA binding capabilities of MBD and ID. The influence of distant disordered domains on MBD properties makes it necessary to consider the NTD-MBD-ID variant as the minimal protein construct for studying dsDNA/chromatin binding properties, while the full-length protein should be considered for transcriptional regulation studies.

LEDGF interacts with the NID of MeCP2 and modulates MeCP2 condensates

“…MeCP2 is an intrinsically disordered protein 32,33 . Characteristically, these proteins lack a stable three-dimensional structure and interact electrostatically with DNA and other proteins.…”

“…In MeCP2 only the MBD is structured, while the ID and TRD are structurally disordered but form a secondary structure upon binding interaction partners and/or DNA 1,34 . We postulated an electrostatic interaction between the ID-TRD and LEDGF which is often the case for structurally disordered proteins 32,33 . Three positively charged regions are present in the TRD and we mutated the four or five positively charged residues to alanines (Figure 5A).…”

The Moonlighting Function of Soybean Disordered Methyl-CpG-Binding Domain 10c Protein

“…The first identified MBD protein, MeCP2, is partially disordered, multifunctional and consists of six domains. The MBD-flanking domains (NTD and ID) are completely disordered and can alter the structure and function of the MBD domain [39]. Soybean GmMBD10c protein belongs to the class 1 MBD protein family and contains a high ratio of disorder-promoting amino acids.…”

Stabilization Effect of Intrinsically Disordered Regions on Multidomain Proteins: The Case of the Methyl-CpG Protein 2, MeCP2

“…It has been reported that both NTD and ID increase the intrinsic structural stability of MBD, as observed by thermal unfolding experiments: at pH 7, the unfolding temperature T m is 38.4 • C and 46.2 • C for MBD and NTD-MBD-ID, respectively, and the unfolding enthalpy ∆H m is 38 kcal/mol and 46 kcal/mol for MBD and NTD-MBD-ID, respectively [34]. All other domains also have a minor stabilizing effect on MBD [35]. These unfolding experiments were undertaken by following the intrinsic fluorescence emission of the single tryptophan located at the MBD (W104), and consequently they accurately reflect the thermal stability of MBD.…”