Structure of the human histone chaperone FACT Spt16 N-terminal domain

The histone chaperone FACT plays an important role in facilitating nucleosome assembly and disassembly during transcription. FACT is a heterodimeric complex consisting of Spt16 and SSRP1. The N-terminal domain of Spt16 resembles an inactive aminopeptidase. How this domain contributes to the histone chaperone activity of FACT remains elusive. Here, the crystal structure of the N-terminal domain (NTD) of human Spt16 is reported at a resolution of 1.84 Å. The structure adopts an aminopeptidase-like fold similar to those of theSaccharomyces cerevisiaeandSchizosaccharomyces pombeSpt16 NTDs. Isothermal titration calorimetry analyses show that human Spt16 NTD binds histones H3/H4 with low-micromolar affinity, suggesting that Spt16 NTD may contribute to histone binding in the FACT complex. Surface-residue conservation and electrostatic analysis reveal a conserved acidic patch that may be involved in histone binding.

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Faculty Opinions recommendation of The crystal structure of nucleoplasmin-core: implications for histone binding and nucleosome assembly.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1002278.46554 ◽

2002 ◽

Author(s):

Song Tan

Keyword(s):

Crystal Structure ◽

Nucleosome Assembly ◽

Histone Binding

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Faculty Opinions recommendation of The crystal structure of nucleoplasmin-core: implications for histone binding and nucleosome assembly.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1002278.27706 ◽

2001 ◽

Author(s):

Elena Conti

Keyword(s):

Crystal Structure ◽

Nucleosome Assembly ◽

Histone Binding

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Structural and histone binding studies of the chromo barrel domain of TIP60

10.1101/257485 ◽

2018 ◽

Author(s):

Yuzhe Zhang ◽

Ming Lei ◽

Xiao Liang ◽

Peter Loppnau ◽

Yanjun Li ◽

...

Keyword(s):

Crystal Structure ◽

Peptide Binding ◽

Side Chain ◽

Titration Calorimetry ◽

Binding Studies ◽

Histone Tails ◽

Histone Binding ◽

Cellular Processes ◽

Peptide Binding Site ◽

Acetyltransferase Domain

AbstractTIP60 consists of an N-terminal chromo barrel domain (TIP60-CB) and a C-terminal acetyltransferase domain and acetylates histone and non-histone proteins within diverse cellular processes. Whereas the TIP60-CB is thought to recognize histone tails, molecular details of this interaction remain unclear. Here we attempted a quantitative analysis of the interaction between the TIP60-CB and histone peptides, but did not observe any binding through either fluorescence polarization or isothermal titration calorimetry. We solved a crystal structure of the TIP60-CB alone. Analysis of the crystal structure demonstrates a putative peptide binding site that may be occluded by the basic side chain of a residue in a unique β hairpin between the two N-terminal strands of the β barrel.

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Crystal structure of SARS-CoV-2 Orf9b in complex with human TOM70 suggests unusual virus-host interactions

Nature Communications ◽

10.1038/s41467-021-23118-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Xiaopan Gao ◽

Kaixiang Zhu ◽

Bo Qin ◽

Vincent Olieric ◽

Meitian Wang ◽

...

Keyword(s):

Crystal Structure ◽

Heat Shock Protein 90 ◽

Helical Conformation ◽

Titration Calorimetry ◽

C Peptide ◽

Host Interactions ◽

Terminal Domain ◽

Host Interaction ◽

Endothermic Process ◽

Interferon Responses

AbstractAlthough the accessory proteins are considered non-essential for coronavirus replication, accumulating evidences demonstrate they are critical to virus-host interaction and pathogenesis. Orf9b is a unique accessory protein of SARS-CoV-2 and SARS-CoV. It is implicated in immune evasion by targeting mitochondria, where it associates with the versatile adapter TOM70. Here, we determined the crystal structure of SARS-CoV-2 orf9b in complex with the cytosolic segment of human TOM70 to 2.2 Å. A central portion of orf9b occupies the deep pocket in the TOM70 C-terminal domain (CTD) and adopts a helical conformation strikingly different from the β-sheet-rich structure of the orf9b homodimer. Interactions between orf9b and TOM70 CTD are primarily hydrophobic and distinct from the electrostatic interaction between the heat shock protein 90 (Hsp90) EEVD motif and the TOM70 N-terminal domain (NTD). Using isothermal titration calorimetry (ITC), we demonstrated that the orf9b dimer does not bind TOM70, but a synthetic peptide harboring a segment of orf9b (denoted C-peptide) binds TOM70 with nanomolar KD. While the interaction between C-peptide and TOM70 CTD is an endothermic process, the interaction between Hsp90 EEVD and TOM70 NTD is exothermic, which underscores the distinct binding mechanisms at NTD and CTD pockets. Strikingly, the binding affinity of Hsp90 EEVD motif to TOM70 NTD is reduced by ~29-fold when orf9b occupies the pocket of TOM70 CTD, supporting the hypothesis that orf9b allosterically inhibits the Hsp90/TOM70 interaction. Our findings shed light on the mechanism underlying SARS-CoV-2 orf9b mediated suppression of interferon responses.

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Faculty Opinions recommendation of The crystal structure of nucleoplasmin-core: implications for histone binding and nucleosome assembly.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1002278.20504 ◽

2001 ◽

Author(s):

Robert Dutnall

Keyword(s):

Crystal Structure ◽

Nucleosome Assembly ◽

Histone Binding

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AtFKBP53: a chimeric histone chaperone with functional nucleoplasmin and PPIase domains

Nucleic Acids Research ◽

10.1093/nar/gkz1153 ◽

2019 ◽

Vol 48 (3) ◽

pp. 1531-1550

Author(s):

Ajit Kumar Singh ◽

Aritreyee Datta ◽

Chacko Jobichen ◽

Sheng Luan ◽

Dileep Vasudevan

Keyword(s):

Binding Sites ◽

Histone H3 ◽

Histone Chaperone ◽

Nucleosome Assembly ◽

Ppiase Activity ◽

Terminal Domain ◽

C Terminus ◽

Ppiase Domain ◽

Fk506 Binding Proteins ◽

Discrete Complex

Abstract FKBP53 is one of the seven multi-domain FK506-binding proteins present in Arabidopsis thaliana, and it is known to get targeted to the nucleus. It has a conserved PPIase domain at the C-terminus and a highly charged N-terminal stretch, which has been reported to bind to histone H3 and perform the function of a histone chaperone. To better understand the molecular details of this PPIase with histone chaperoning activity, we have solved the crystal structures of its terminal domains and functionally characterized them. The C-terminal domain showed strong PPIase activity, no role in histone chaperoning and revealed a monomeric five-beta palm-like fold that wrapped over a helix, typical of an FK506-binding domain. The N-terminal domain had a pentameric nucleoplasmin-fold; making this the first report of a plant nucleoplasmin structure. Further characterization revealed the N-terminal nucleoplasmin domain to interact with H2A/H2B and H3/H4 histone oligomers, individually, as well as simultaneously, suggesting two different binding sites for H2A/H2B and H3/H4. The pentameric domain assists nucleosome assembly and forms a discrete complex with pre-formed nucleosomes; wherein two pentamers bind to a nucleosome.

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Structural Characterization of Arabidopsis thaliana NAP1-Related Protein 2 (AtNRP2) and Comparison with its Homolog AtNRP1

Molecules ◽

10.3390/molecules24122258 ◽

2019 ◽

Vol 24 (12) ◽

pp. 2258 ◽

Cited By ~ 4

Author(s):

Kumar ◽

Kumar Singh ◽

Chandrakant Bobde ◽

Vasudevan

Keyword(s):

Crystal Structure ◽

Arabidopsis Thaliana ◽

Titration Calorimetry ◽

Histone Chaperones ◽

Nucleosome Assembly ◽

Mobility Shift ◽

Mobility Shift Assay ◽

The Stability ◽

Structural Architecture ◽

And Function

Nucleosome Assembly Protein (NAP) is a highly conserved family of histone chaperones present in yeast, animals, and plants. Unlike other organisms, plants possess an additional class of proteins in its NAP family, known as the NAP1-related proteins or NRP. Arabidopsis thaliana possesses two NRP isoforms, namely AtNRP1 and AtNRP2, that share 87% sequence identity. Both AtNRP1 and AtNRP2 get expressed in all the plant tissues. Most works in the past, including structural studies, have focused on AtNRP1. We wanted to do a comparative study of the two proteins to find why the plant would have two very similar proteins and whether there is any difference between the two for their structure and function as histone chaperones. Here we report the crystal structure of AtNRP2 and a comparative analysis of its structural architecture with other NAP family proteins. The crystal structure of AtNRP2 shows it to be a homodimer, with its fold similar to that of other structurally characterized NAP family proteins. Although AtNRP1 and AtNRP2 have a similar fold, upon structural superposition, we find an offset in the dimerization helix of the two proteins. We evaluated the stability, oligomerization status, and histone chaperoning properties of the two proteins, for a comparison. The thermal melting experiments suggest that AtNRP2 is more stable than AtNRP1 at higher temperatures. In addition, electrophoretic mobility shift assay and isothermal titration calorimetry experiments suggest histone binding ability of AtNRP2 is higher than that of AtNRP1. Overall, these results provide insights about the specific function and relevance of AtNRP2 in plants through structural and biophysical studies.

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