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Recognition of asymmetrically dimethylated arginine by TDRD3.

Sikorsky T, Hobor F, Krizanova E, Pasulka J, Kubicek K, Stefl R - Nucleic Acids Res. (2012)

Bottom Line: The structure and mutational analysis provide a molecular basis for how TDRD3 recognizes the aDMA mark.The unique aromatic cavity of the TDRD3 Tudor domain with a tyrosine in position 566 creates a selectivity filter for the aDMA residue.Our work contributes to the understanding of substrate selectivity rules of the Tudor aromatic cavity, which is an important structural motif for reading of methylation marks.

View Article: PubMed Central - PubMed

Affiliation: CEITEC-Central European Institute of Technology, Masaryk University, CZ-62500 Brno, Czech Republic.

ABSTRACT
Asymmetric dimethylarginine (aDMA) marks are placed on histones and the C-terminal domain (CTD) of RNA Polymerase II (RNAP II) and serve as a signal for recruitment of appropriate transcription and processing factors in coordination with transcription cycle. In contrast to other Tudor domain-containing proteins, Tudor domain-containing protein 3 (TDRD3) associates selectively with the aDMA marks but not with other methylarginine motifs. Here, we report the solution structure of the Tudor domain of TDRD3 bound to the asymmetrically dimethylated CTD. The structure and mutational analysis provide a molecular basis for how TDRD3 recognizes the aDMA mark. The unique aromatic cavity of the TDRD3 Tudor domain with a tyrosine in position 566 creates a selectivity filter for the aDMA residue. Our work contributes to the understanding of substrate selectivity rules of the Tudor aromatic cavity, which is an important structural motif for reading of methylation marks.

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Recognition of aDMA by the TDRD3 Tudor. (A) Sequence alignment of the human TDRD3 Tudor domain with other dimethylarginine binding Tudor domains of SMN (Homo sapiens), SPF30 (H. sapiens), SND1 (H. sapiens), and the 11th Tudor domain of Tud (Drosophila melanogaster; Tud11). Residues forming an aromatic cavity are highlighted in green squares, variable residues are shown in red. The β-sheet regions (β1, β2, β3 and β4) of human TDRD3 Tudor are shown with blue arrows. (B) Bar plot of the NMR-derived association constants (Ka) of various TDRD3 mutants with the aDMA-CTD peptide (blue) and sDMA-CTD peptide (red) in a logarithmic scale. Asterisk indicates that the binding constant could not be determined. (C) Bar plot showing decomposed interactions’ energies between the aromatic cavity and aDMA calculated by SAPT. The exchange-repulsion (blue bar) accounts for an interaction caused by tunneling of the electrons between interacting systems and electron−electron repulsion due to the Pauli exclusion principle. The induction interaction (yellow bar) is a second-order energy contribution, which originates from mutual distortion of electron density distribution of interacting molecules. The dispersion interaction (green) arises from the correlated electron fluctuations in the interacting molecules (48). (D) Pyramidalization of the aDMA amino group as predicted by DFT-D theory. The hydrogen bond (2.7 Å) that is responsible for aDMA recognition is shown with yellow dotted line. Only heavy atoms and non-polar hydrogens are shown.
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gks929-F3: Recognition of aDMA by the TDRD3 Tudor. (A) Sequence alignment of the human TDRD3 Tudor domain with other dimethylarginine binding Tudor domains of SMN (Homo sapiens), SPF30 (H. sapiens), SND1 (H. sapiens), and the 11th Tudor domain of Tud (Drosophila melanogaster; Tud11). Residues forming an aromatic cavity are highlighted in green squares, variable residues are shown in red. The β-sheet regions (β1, β2, β3 and β4) of human TDRD3 Tudor are shown with blue arrows. (B) Bar plot of the NMR-derived association constants (Ka) of various TDRD3 mutants with the aDMA-CTD peptide (blue) and sDMA-CTD peptide (red) in a logarithmic scale. Asterisk indicates that the binding constant could not be determined. (C) Bar plot showing decomposed interactions’ energies between the aromatic cavity and aDMA calculated by SAPT. The exchange-repulsion (blue bar) accounts for an interaction caused by tunneling of the electrons between interacting systems and electron−electron repulsion due to the Pauli exclusion principle. The induction interaction (yellow bar) is a second-order energy contribution, which originates from mutual distortion of electron density distribution of interacting molecules. The dispersion interaction (green) arises from the correlated electron fluctuations in the interacting molecules (48). (D) Pyramidalization of the aDMA amino group as predicted by DFT-D theory. The hydrogen bond (2.7 Å) that is responsible for aDMA recognition is shown with yellow dotted line. Only heavy atoms and non-polar hydrogens are shown.

Mentions: It was shown that substitution of any of the four aromatic residues of the cavity with a non-aromatic amino acid, abrogates dimethylarginine binding (26). In the present study, we have investigated the effect of aromatic substitutions of the least conserved residue within the aromatic cavity (Figures 2C and 3A). In TDRD3, Y566 is a unique residue, whereas SMN, SPF30, SND1 and Tur11 contain tryptophan or phenylalanine in this position. Y566F substitution diminishes binding to aDMA-CTD and it does not increase binding affinity to sDMA-CTD (Figure 3B and Supplementary Figure S5). On the other hand, Y566W substitution promotes complex formation with sDMA-CTD, yet it has similar binding affinity to aDMA-CTD as the wild-type protein. This indicates that both phenylalanine and tryptophan substitutions at position Y566 abrogate TDRD3 selectivity for aDMA- against sDMA-CTD peptides. Residue Y566 is thus the key element, which determines the specificity of TDRD3 toward aDMA-containing peptides.Figure 3.


Recognition of asymmetrically dimethylated arginine by TDRD3.

Sikorsky T, Hobor F, Krizanova E, Pasulka J, Kubicek K, Stefl R - Nucleic Acids Res. (2012)

Recognition of aDMA by the TDRD3 Tudor. (A) Sequence alignment of the human TDRD3 Tudor domain with other dimethylarginine binding Tudor domains of SMN (Homo sapiens), SPF30 (H. sapiens), SND1 (H. sapiens), and the 11th Tudor domain of Tud (Drosophila melanogaster; Tud11). Residues forming an aromatic cavity are highlighted in green squares, variable residues are shown in red. The β-sheet regions (β1, β2, β3 and β4) of human TDRD3 Tudor are shown with blue arrows. (B) Bar plot of the NMR-derived association constants (Ka) of various TDRD3 mutants with the aDMA-CTD peptide (blue) and sDMA-CTD peptide (red) in a logarithmic scale. Asterisk indicates that the binding constant could not be determined. (C) Bar plot showing decomposed interactions’ energies between the aromatic cavity and aDMA calculated by SAPT. The exchange-repulsion (blue bar) accounts for an interaction caused by tunneling of the electrons between interacting systems and electron−electron repulsion due to the Pauli exclusion principle. The induction interaction (yellow bar) is a second-order energy contribution, which originates from mutual distortion of electron density distribution of interacting molecules. The dispersion interaction (green) arises from the correlated electron fluctuations in the interacting molecules (48). (D) Pyramidalization of the aDMA amino group as predicted by DFT-D theory. The hydrogen bond (2.7 Å) that is responsible for aDMA recognition is shown with yellow dotted line. Only heavy atoms and non-polar hydrogens are shown.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
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gks929-F3: Recognition of aDMA by the TDRD3 Tudor. (A) Sequence alignment of the human TDRD3 Tudor domain with other dimethylarginine binding Tudor domains of SMN (Homo sapiens), SPF30 (H. sapiens), SND1 (H. sapiens), and the 11th Tudor domain of Tud (Drosophila melanogaster; Tud11). Residues forming an aromatic cavity are highlighted in green squares, variable residues are shown in red. The β-sheet regions (β1, β2, β3 and β4) of human TDRD3 Tudor are shown with blue arrows. (B) Bar plot of the NMR-derived association constants (Ka) of various TDRD3 mutants with the aDMA-CTD peptide (blue) and sDMA-CTD peptide (red) in a logarithmic scale. Asterisk indicates that the binding constant could not be determined. (C) Bar plot showing decomposed interactions’ energies between the aromatic cavity and aDMA calculated by SAPT. The exchange-repulsion (blue bar) accounts for an interaction caused by tunneling of the electrons between interacting systems and electron−electron repulsion due to the Pauli exclusion principle. The induction interaction (yellow bar) is a second-order energy contribution, which originates from mutual distortion of electron density distribution of interacting molecules. The dispersion interaction (green) arises from the correlated electron fluctuations in the interacting molecules (48). (D) Pyramidalization of the aDMA amino group as predicted by DFT-D theory. The hydrogen bond (2.7 Å) that is responsible for aDMA recognition is shown with yellow dotted line. Only heavy atoms and non-polar hydrogens are shown.
Mentions: It was shown that substitution of any of the four aromatic residues of the cavity with a non-aromatic amino acid, abrogates dimethylarginine binding (26). In the present study, we have investigated the effect of aromatic substitutions of the least conserved residue within the aromatic cavity (Figures 2C and 3A). In TDRD3, Y566 is a unique residue, whereas SMN, SPF30, SND1 and Tur11 contain tryptophan or phenylalanine in this position. Y566F substitution diminishes binding to aDMA-CTD and it does not increase binding affinity to sDMA-CTD (Figure 3B and Supplementary Figure S5). On the other hand, Y566W substitution promotes complex formation with sDMA-CTD, yet it has similar binding affinity to aDMA-CTD as the wild-type protein. This indicates that both phenylalanine and tryptophan substitutions at position Y566 abrogate TDRD3 selectivity for aDMA- against sDMA-CTD peptides. Residue Y566 is thus the key element, which determines the specificity of TDRD3 toward aDMA-containing peptides.Figure 3.

Bottom Line: The structure and mutational analysis provide a molecular basis for how TDRD3 recognizes the aDMA mark.The unique aromatic cavity of the TDRD3 Tudor domain with a tyrosine in position 566 creates a selectivity filter for the aDMA residue.Our work contributes to the understanding of substrate selectivity rules of the Tudor aromatic cavity, which is an important structural motif for reading of methylation marks.

View Article: PubMed Central - PubMed

Affiliation: CEITEC-Central European Institute of Technology, Masaryk University, CZ-62500 Brno, Czech Republic.

ABSTRACT
Asymmetric dimethylarginine (aDMA) marks are placed on histones and the C-terminal domain (CTD) of RNA Polymerase II (RNAP II) and serve as a signal for recruitment of appropriate transcription and processing factors in coordination with transcription cycle. In contrast to other Tudor domain-containing proteins, Tudor domain-containing protein 3 (TDRD3) associates selectively with the aDMA marks but not with other methylarginine motifs. Here, we report the solution structure of the Tudor domain of TDRD3 bound to the asymmetrically dimethylated CTD. The structure and mutational analysis provide a molecular basis for how TDRD3 recognizes the aDMA mark. The unique aromatic cavity of the TDRD3 Tudor domain with a tyrosine in position 566 creates a selectivity filter for the aDMA residue. Our work contributes to the understanding of substrate selectivity rules of the Tudor aromatic cavity, which is an important structural motif for reading of methylation marks.

Show MeSH
Related in: MedlinePlus