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Structure of the JmjC domain-containing protein NO66 complexed with ribosomal protein Rpl8.

Wang C, Zhang Q, Hang T, Tao Y, Ma X, Wu M, Zhang X, Zang J - Acta Crystallogr. D Biol. Crystallogr. (2015)

Bottom Line: Based on the results of structural and biochemical analyses, the consensus sequence motif NHXH recognized by NO66 was confirmed.When binding to substrate, the relative positions of each subunit in the NO66 tetramer shift.Oligomerization may facilitate the motion of each subunit in the NO66 tetramer and affect the catalytic activity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, Collaborative Innovation Center of Chemistry for Life Science, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230026, People's Republic of China.

ABSTRACT
The JmjC domain-containing proteins belong to a large family of oxygenases possessing distinct substrate specificities which are involved in the regulation of different biological processes, such as gene transcription, RNA processing and translation. Nucleolar protein 66 (NO66) is a JmjC domain-containing protein which has been reported to be a histone demethylase and a ribosome protein 8 (Rpl8) hydroxylase. The present biochemical study confirmed the hydroxylase activity of NO66 and showed that oligomerization is required for NO66 to efficiently catalyze the hydroxylation of Rpl8. The structures of NO66(176-C) complexed with Rpl8(204-224) in a tetrameric form and of the mutant protein M2 in a dimeric form were solved. Based on the results of structural and biochemical analyses, the consensus sequence motif NHXH recognized by NO66 was confirmed. Several potential substrates of NO66 were found by a BLAST search according to the consensus sequence motif. When binding to substrate, the relative positions of each subunit in the NO66 tetramer shift. Oligomerization may facilitate the motion of each subunit in the NO66 tetramer and affect the catalytic activity.

No MeSH data available.


Specific recognition of Rpl8 by NO66. (a) A surface representation of the active site of NO66 is shown. Positively charged, negatively charged and neutral areas are shown in blue, red and white, respectively. The Rpl8204–224 peptide adopts a U-shaped conformation in the active site of NO66. (b) A detailed view of the interaction network of NO66 with Rpl8204–224 and NOG. Rpl8204–224 and the amino-acid residues of NO66 interacting with Rpl8204–224 are shown as stick models. Ni2+ and water molecules are shown as spheres. NO66, Rpl8204–224, NOG, Ni2+ and water molecules are coloured grey, yellow, green, magenta and red, respectively. (c) His216 is the hydroxylation site in Rpl8 catalyzed by NO66. The distances between the Cβ atoms of His216 and His218 and the Ni2+ ion in the active site are shown.
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fig3: Specific recognition of Rpl8 by NO66. (a) A surface representation of the active site of NO66 is shown. Positively charged, negatively charged and neutral areas are shown in blue, red and white, respectively. The Rpl8204–224 peptide adopts a U-shaped conformation in the active site of NO66. (b) A detailed view of the interaction network of NO66 with Rpl8204–224 and NOG. Rpl8204–224 and the amino-acid residues of NO66 interacting with Rpl8204–224 are shown as stick models. Ni2+ and water molecules are shown as spheres. NO66, Rpl8204–224, NOG, Ni2+ and water molecules are coloured grey, yellow, green, magenta and red, respectively. (c) His216 is the hydroxylation site in Rpl8 catalyzed by NO66. The distances between the Cβ atoms of His216 and His218 and the Ni2+ ion in the active site are shown.

Mentions: In the complex structure of NO66176–C and Rpl8204–224 only residues 212–222 of Rpl8 are visible in the electron-density map. The other regions of this peptide are invisible, possibly because of high flexibility. From the complex structure, it is apparent that the Rpl8204–224 peptide bound to NO66176–C in a cleft located in the centre of the JmjC domain (Fig. 3 ▸a). The overall surface area of NO66 covered by the bound peptide is about 659 Å2. The Rpl8204–224 peptide adopts a U-shaped conformation and penetrates deeply into a cleft (Fig. 3 ▸a). Residue Gln217 points to the outside of the binding pocket, leading to a bulge shape formed in the middle of the U-shaped Rpl8204–224 peptide. This bulge-shaped structure is stabilized by a hydrogen bond formed between the backbone carbonyl group of Gly217 and the backbone amide group of Gly220 of Rpl8204–224 (Fig. 2 ▸b).


Structure of the JmjC domain-containing protein NO66 complexed with ribosomal protein Rpl8.

Wang C, Zhang Q, Hang T, Tao Y, Ma X, Wu M, Zhang X, Zang J - Acta Crystallogr. D Biol. Crystallogr. (2015)

Specific recognition of Rpl8 by NO66. (a) A surface representation of the active site of NO66 is shown. Positively charged, negatively charged and neutral areas are shown in blue, red and white, respectively. The Rpl8204–224 peptide adopts a U-shaped conformation in the active site of NO66. (b) A detailed view of the interaction network of NO66 with Rpl8204–224 and NOG. Rpl8204–224 and the amino-acid residues of NO66 interacting with Rpl8204–224 are shown as stick models. Ni2+ and water molecules are shown as spheres. NO66, Rpl8204–224, NOG, Ni2+ and water molecules are coloured grey, yellow, green, magenta and red, respectively. (c) His216 is the hydroxylation site in Rpl8 catalyzed by NO66. The distances between the Cβ atoms of His216 and His218 and the Ni2+ ion in the active site are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556315&req=5

fig3: Specific recognition of Rpl8 by NO66. (a) A surface representation of the active site of NO66 is shown. Positively charged, negatively charged and neutral areas are shown in blue, red and white, respectively. The Rpl8204–224 peptide adopts a U-shaped conformation in the active site of NO66. (b) A detailed view of the interaction network of NO66 with Rpl8204–224 and NOG. Rpl8204–224 and the amino-acid residues of NO66 interacting with Rpl8204–224 are shown as stick models. Ni2+ and water molecules are shown as spheres. NO66, Rpl8204–224, NOG, Ni2+ and water molecules are coloured grey, yellow, green, magenta and red, respectively. (c) His216 is the hydroxylation site in Rpl8 catalyzed by NO66. The distances between the Cβ atoms of His216 and His218 and the Ni2+ ion in the active site are shown.
Mentions: In the complex structure of NO66176–C and Rpl8204–224 only residues 212–222 of Rpl8 are visible in the electron-density map. The other regions of this peptide are invisible, possibly because of high flexibility. From the complex structure, it is apparent that the Rpl8204–224 peptide bound to NO66176–C in a cleft located in the centre of the JmjC domain (Fig. 3 ▸a). The overall surface area of NO66 covered by the bound peptide is about 659 Å2. The Rpl8204–224 peptide adopts a U-shaped conformation and penetrates deeply into a cleft (Fig. 3 ▸a). Residue Gln217 points to the outside of the binding pocket, leading to a bulge shape formed in the middle of the U-shaped Rpl8204–224 peptide. This bulge-shaped structure is stabilized by a hydrogen bond formed between the backbone carbonyl group of Gly217 and the backbone amide group of Gly220 of Rpl8204–224 (Fig. 2 ▸b).

Bottom Line: Based on the results of structural and biochemical analyses, the consensus sequence motif NHXH recognized by NO66 was confirmed.When binding to substrate, the relative positions of each subunit in the NO66 tetramer shift.Oligomerization may facilitate the motion of each subunit in the NO66 tetramer and affect the catalytic activity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, Collaborative Innovation Center of Chemistry for Life Science, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui 230026, People's Republic of China.

ABSTRACT
The JmjC domain-containing proteins belong to a large family of oxygenases possessing distinct substrate specificities which are involved in the regulation of different biological processes, such as gene transcription, RNA processing and translation. Nucleolar protein 66 (NO66) is a JmjC domain-containing protein which has been reported to be a histone demethylase and a ribosome protein 8 (Rpl8) hydroxylase. The present biochemical study confirmed the hydroxylase activity of NO66 and showed that oligomerization is required for NO66 to efficiently catalyze the hydroxylation of Rpl8. The structures of NO66(176-C) complexed with Rpl8(204-224) in a tetrameric form and of the mutant protein M2 in a dimeric form were solved. Based on the results of structural and biochemical analyses, the consensus sequence motif NHXH recognized by NO66 was confirmed. Several potential substrates of NO66 were found by a BLAST search according to the consensus sequence motif. When binding to substrate, the relative positions of each subunit in the NO66 tetramer shift. Oligomerization may facilitate the motion of each subunit in the NO66 tetramer and affect the catalytic activity.

No MeSH data available.