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Intra- and inter-nucleosomal interactions of the histone H4 tail revealed with a human nucleosome core particle with genetically-incorporated H4 tetra-acetylation.

Wakamori M, Fujii Y, Suka N, Shirouzu M, Sakamoto K, Umehara T, Yokoyama S - Sci Rep (2015)

Bottom Line: However, the B-factors were significantly increased for the peripheral DNAs near the N-terminal tail of the intra- or inter-nucleosomal H4.In contrast, the B-factors were negligibly affected by the H4 tetra-acetylation in histone core residues, including those composing the acidic patch, and at H4-R23, which interacts with the acidic patch of the neighboring NCP.The present study revealed that the H4 tetra-acetylation impairs NCP self-association by changing the interactions of the H4 tail with DNA, and is the first demonstration of crystallization quality NCPs reconstituted with genuine PTMs.

View Article: PubMed Central - PubMed

Affiliation: RIKEN Systems and Structural Biology Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.

ABSTRACT
Post-translational modifications (PTMs) of histones, such as lysine acetylation of the N-terminal tails, play crucial roles in controlling gene expression. Due to the difficulty in reconstituting site-specifically acetylated nucleosomes with crystallization quality, structural analyses of histone acetylation are currently performed using synthesized tail peptides. Through engineering of the genetic code, translation termination, and cell-free protein synthesis, we reconstituted human H4-mono- to tetra-acetylated nucleosome core particles (NCPs), and solved the crystal structures of the H4-K5/K8/K12/K16-tetra-acetylated NCP and unmodified NCP at 2.4 Å and 2.2 Å resolutions, respectively. The structure of the H4-tetra-acetylated NCP resembled that of the unmodified NCP, and the DNA wrapped the histone octamer as precisely as in the unmodified NCP. However, the B-factors were significantly increased for the peripheral DNAs near the N-terminal tail of the intra- or inter-nucleosomal H4. In contrast, the B-factors were negligibly affected by the H4 tetra-acetylation in histone core residues, including those composing the acidic patch, and at H4-R23, which interacts with the acidic patch of the neighboring NCP. The present study revealed that the H4 tetra-acetylation impairs NCP self-association by changing the interactions of the H4 tail with DNA, and is the first demonstration of crystallization quality NCPs reconstituted with genuine PTMs.

No MeSH data available.


Related in: MedlinePlus

Representation of B-factor-increased nucleosomal DNA regions by the H4 tetra-acetylation.(a) Intra-nucleosomal contacts by the molecule-B H4. The DNA chains are color-coded by the difference in the B-factors between the H4-tetra-acetylated and unmodified NCPs. The color bar represents the B-factor difference from 0 Å2 (blue) to 30 Å2 (red). (b) Intra-nucleosomal contacts by the molecule-F H4. (c) Inter-nucleosomal contacts by the molecule-B H4.
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f4: Representation of B-factor-increased nucleosomal DNA regions by the H4 tetra-acetylation.(a) Intra-nucleosomal contacts by the molecule-B H4. The DNA chains are color-coded by the difference in the B-factors between the H4-tetra-acetylated and unmodified NCPs. The color bar represents the B-factor difference from 0 Å2 (blue) to 30 Å2 (red). (b) Intra-nucleosomal contacts by the molecule-F H4. (c) Inter-nucleosomal contacts by the molecule-B H4.

Mentions: In contrast, the outward DNA around SHL +2 is near the visible N-terminal residue of the intra-nucleosomal molecule-F H4, but its B-factor differences are comparatively modest (dashed circles in Fig. 3a,b). Close-up views of the nucleosomal DNAs near the H4 tail of molecule B or F are shown in Fig. 4a–c. The positions of the DNA backbone with large B-factor differences around 30 Å2 are color-coded in red. As residues 1–20 and 1–15 of molecules B and F, respectively, are disordered in the crystals, we roughly estimated the distances from the visible N-terminal residues, V21 of molecule B and K16ac of molecule F, to one of the acetyllysine-incorporated residues, K12ac, using the average inter-residue distance of approximately 3.5 Å in a typical β strand, as 31.5 Å (9 residues) and 14 Å (4 residues), respectively. These distances are indicated as the radii of the circles. From this representation, we found that although the electron densities corresponding to the N-terminal 20 residues are disordered in the molecule-B H4, the H4-tetra-acetylation of the molecule-B H4 seems to locally affect the B-factors of both the intra-nucleosomal DNA (Fig. 4a) and inter-nucleosomal DNA (Fig. 4c). However, the H4-tetra-acetylation of the molecule-F H4 affected the B-factors of the nearby intra-nucleosomal DNA to a lower extent (Fig. 4b; see also dashed circles in Fig. 3a,b). Finally, we averaged the B-factors of all nucleotides, except the significantly affected ones, for each of the DNA chains, and confirmed that there was no statistically significant effect of the H4-tetra-acetylation.


Intra- and inter-nucleosomal interactions of the histone H4 tail revealed with a human nucleosome core particle with genetically-incorporated H4 tetra-acetylation.

Wakamori M, Fujii Y, Suka N, Shirouzu M, Sakamoto K, Umehara T, Yokoyama S - Sci Rep (2015)

Representation of B-factor-increased nucleosomal DNA regions by the H4 tetra-acetylation.(a) Intra-nucleosomal contacts by the molecule-B H4. The DNA chains are color-coded by the difference in the B-factors between the H4-tetra-acetylated and unmodified NCPs. The color bar represents the B-factor difference from 0 Å2 (blue) to 30 Å2 (red). (b) Intra-nucleosomal contacts by the molecule-F H4. (c) Inter-nucleosomal contacts by the molecule-B H4.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4660432&req=5

f4: Representation of B-factor-increased nucleosomal DNA regions by the H4 tetra-acetylation.(a) Intra-nucleosomal contacts by the molecule-B H4. The DNA chains are color-coded by the difference in the B-factors between the H4-tetra-acetylated and unmodified NCPs. The color bar represents the B-factor difference from 0 Å2 (blue) to 30 Å2 (red). (b) Intra-nucleosomal contacts by the molecule-F H4. (c) Inter-nucleosomal contacts by the molecule-B H4.
Mentions: In contrast, the outward DNA around SHL +2 is near the visible N-terminal residue of the intra-nucleosomal molecule-F H4, but its B-factor differences are comparatively modest (dashed circles in Fig. 3a,b). Close-up views of the nucleosomal DNAs near the H4 tail of molecule B or F are shown in Fig. 4a–c. The positions of the DNA backbone with large B-factor differences around 30 Å2 are color-coded in red. As residues 1–20 and 1–15 of molecules B and F, respectively, are disordered in the crystals, we roughly estimated the distances from the visible N-terminal residues, V21 of molecule B and K16ac of molecule F, to one of the acetyllysine-incorporated residues, K12ac, using the average inter-residue distance of approximately 3.5 Å in a typical β strand, as 31.5 Å (9 residues) and 14 Å (4 residues), respectively. These distances are indicated as the radii of the circles. From this representation, we found that although the electron densities corresponding to the N-terminal 20 residues are disordered in the molecule-B H4, the H4-tetra-acetylation of the molecule-B H4 seems to locally affect the B-factors of both the intra-nucleosomal DNA (Fig. 4a) and inter-nucleosomal DNA (Fig. 4c). However, the H4-tetra-acetylation of the molecule-F H4 affected the B-factors of the nearby intra-nucleosomal DNA to a lower extent (Fig. 4b; see also dashed circles in Fig. 3a,b). Finally, we averaged the B-factors of all nucleotides, except the significantly affected ones, for each of the DNA chains, and confirmed that there was no statistically significant effect of the H4-tetra-acetylation.

Bottom Line: However, the B-factors were significantly increased for the peripheral DNAs near the N-terminal tail of the intra- or inter-nucleosomal H4.In contrast, the B-factors were negligibly affected by the H4 tetra-acetylation in histone core residues, including those composing the acidic patch, and at H4-R23, which interacts with the acidic patch of the neighboring NCP.The present study revealed that the H4 tetra-acetylation impairs NCP self-association by changing the interactions of the H4 tail with DNA, and is the first demonstration of crystallization quality NCPs reconstituted with genuine PTMs.

View Article: PubMed Central - PubMed

Affiliation: RIKEN Systems and Structural Biology Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.

ABSTRACT
Post-translational modifications (PTMs) of histones, such as lysine acetylation of the N-terminal tails, play crucial roles in controlling gene expression. Due to the difficulty in reconstituting site-specifically acetylated nucleosomes with crystallization quality, structural analyses of histone acetylation are currently performed using synthesized tail peptides. Through engineering of the genetic code, translation termination, and cell-free protein synthesis, we reconstituted human H4-mono- to tetra-acetylated nucleosome core particles (NCPs), and solved the crystal structures of the H4-K5/K8/K12/K16-tetra-acetylated NCP and unmodified NCP at 2.4 Å and 2.2 Å resolutions, respectively. The structure of the H4-tetra-acetylated NCP resembled that of the unmodified NCP, and the DNA wrapped the histone octamer as precisely as in the unmodified NCP. However, the B-factors were significantly increased for the peripheral DNAs near the N-terminal tail of the intra- or inter-nucleosomal H4. In contrast, the B-factors were negligibly affected by the H4 tetra-acetylation in histone core residues, including those composing the acidic patch, and at H4-R23, which interacts with the acidic patch of the neighboring NCP. The present study revealed that the H4 tetra-acetylation impairs NCP self-association by changing the interactions of the H4 tail with DNA, and is the first demonstration of crystallization quality NCPs reconstituted with genuine PTMs.

No MeSH data available.


Related in: MedlinePlus