Molecular architecture of nucleosome remodeling and deacetylase sub-complexes by integrative structure determination

The Nucleosome Remodeling and Deacetylase (NuRD) complex is a chromatin-modifying assembly that regulates gene expression and DNA damage repair. Despite its importance, limited structural information is available on the complex and a detailed understanding of its mechanism is lacking. We investigated the molecular architecture of three NuRD sub-complexes: MTA1-HDAC1-RBBP4 (MHR), MTA1N-HDAC1-MBD3GATAD2CC (MHM), and MTA1-HDAC1-RBBP4-MBD3-GATAD2 (NuDe) using Bayesian integrative structure determination with IMP (Integrative Modeling Platform), drawing on information from SEC-MALLS, DIA-MS, XLMS, negative stain EM, X-ray crystallography, NMR spectroscopy, secondary structure and homology predictions. The structures were corroborated by independent cryo-EM maps, biochemical assays, and known cancer-associated mutations. Our integrative structure of the 2:2:2 MHM complex shows asymmetric binding of MBD3, whereas our structure of the NuDe complex shows MBD3 localized precisely to a single position distant from the MTA1 dimerization interface. Our models suggest a possible mechanism by which asymmetry is introduced in NuRD, and indicate three previously unrecognized subunit interfaces in NuDe: HDAC1C-MTA1BAH, MTA1BAH-MBD3, and HDAC160-100-MBD3. We observed that a significant number of cancer-associated mutations mapped to protein-protein interfaces in NuDe. Our approach also allows us to localize regions of unknown structure, such as HDAC1C and MBD3IDR, thereby resulting in the most complete structural characterization of these NuRD sub-complexes so far.

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Photosynthetic generation of oxygen

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2008.0047 ◽

2008 ◽

Vol 363 (1504) ◽

pp. 2665-2674 ◽

Cited By ~ 44

Author(s):

James Barber

Keyword(s):

Structural Information ◽

Molecular Architecture ◽

Oxygen Generation ◽

X Ray Crystallography ◽

Biological Studies ◽

Wide Range ◽

Photosynthetic Oxygen ◽

Oxygen Evolving ◽

Opening Up ◽

Protein Environment

The oxygen in the atmosphere is derived from light-driven oxidation of water at a catalytic centre contained within a multi-subunit enzyme known as photosystem II (PSII). PSII is located in the photosynthetic membranes of plants, algae and cyanobacteria and its oxygen-evolving centre (OEC) consists of four manganese ions and a calcium ion surrounded by a highly conserved protein environment. Recently, the structure of PSII was elucidated by X-ray crystallography thus revealing details of the molecular architecture of the OEC. This structural information, coupled with an extensive knowledge base derived from a wide range of biophysical, biochemical and molecular biological studies, has provided a framework for understanding the chemistry of photosynthetic oxygen generation as well as opening up debate about its evolutionary origin.

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X-ray free electron laser: opportunities for drug discovery

Essays in Biochemistry ◽

10.1042/ebc20170031 ◽

2017 ◽

Vol 61 (5) ◽

pp. 529-542 ◽

Cited By ~ 12

Author(s):

Robert K.Y. Cheng ◽

Rafael Abela ◽

Michael Hennig

Keyword(s):

Conformational Changes ◽

Structure Determination ◽

Free Electron ◽

Free Electron Laser ◽

Structural Information ◽

Crystal Sample ◽

X Ray ◽

Drug Candidates ◽

X Ray Crystallography ◽

The Impact

Past decades have shown the impact of structural information derived from complexes of drug candidates with their protein targets to facilitate the discovery of safe and effective medicines. Despite recent developments in single particle cryo-electron microscopy, X-ray crystallography has been the main method to derive structural information. The unique properties of X-ray free electron laser (XFEL) with unmet peak brilliance and beam focus allow X-ray diffraction data recording and successful structure determination from smaller and weaker diffracting crystals shortening timelines in crystal optimization. To further capitalize on the XFEL advantage, innovations in crystal sample delivery for the X-ray experiment, data collection and processing methods are required. This development was a key contributor to serial crystallography allowing structure determination at room temperature yielding physiologically more relevant structures. Adding the time resolution provided by the femtosecond X-ray pulse will enable monitoring and capturing of dynamic processes of ligand binding and associated conformational changes with great impact to the design of candidate drug compounds.

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The Tale of CHD4 in DNA Damage Response and Chemotherapeutic Response

Cancer Research and Cellular Therapeutics ◽

10.31579/2640-1053/052 ◽

2019 ◽

Vol 3 (1) ◽

pp. 01-03

Author(s):

Shiaw-Yih Lin ◽

Jing Zhang ◽

David J.H. Shih

Keyword(s):

Dna Damage ◽

Cancer Progression ◽

Tumor Biology ◽

Stem Cell Differentiation ◽

Chemotherapeutic Agents ◽

Nurd Complex ◽

Independent Manner ◽

Nucleosome Remodeling ◽

Damage Repair ◽

Chemotherapeutic Response

The chromatin remodeling factor chromodomain helicase DNA-binding protein 4 (CHD4) is a core component of the nucleosome remodeling and deacetylase (NuRD) complex. Due to its important role in DNA damage repair, CHD4 has been identified as a key determinant in cancer progression, stem cell differentiation, and T cell and B cell development. Accumulating evidence has revealed that CHD4 can function in NuRD dependent and independent manner in response to DNA damage. Mutations of CHD4 have been shown to diminish its functions, which indicates that interpretation of its mutations may provide tangible benefit for patients. The expression of CHD4 play a dual role in sensitizing cancer cells to chemotherapeutic agents, which provides new insights into the contribution of CHD4 to tumor biology and new therapeutic avenues.

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S-SAD phasing of monoclinic histidine kinase fromBrucella abortuscombining data from multiple crystals and orientations: an example of data-collection strategy anda posteriorianalysis of different data combinations

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s1399004715007622 ◽

2015 ◽

Vol 71 (7) ◽

pp. 1433-1443 ◽

Cited By ~ 8

Author(s):

Sebastián Klinke ◽

Nicolas Foos ◽

Jimena J. Rinaldi ◽

Gastón Paris ◽

Fernando A. Goldbaum ◽

...

Keyword(s):

Data Collection ◽

Structure Determination ◽

Histidine Kinase ◽

Structural Information ◽

Data Sets ◽

X Rays ◽

X Ray Crystallography ◽

Data Collection Strategy ◽

Sad Phasing ◽

The Impact

The histidine kinase (HK) domain belonging to the light–oxygen–voltage histidine kinase (LOV-HK) fromBrucella abortusis a member of the HWE family, for which no structural information is available, and has low sequence identity (20%) to the closest HK present in the PDB. The `off-edge' S-SAD method in macromolecular X-ray crystallography was used to solve the structure of the HK domain from LOV-HK at low resolution from crystals in a low-symmetry space group (P21) and with four copies in the asymmetric unit (∼108 kDa). Data were collected both from multiple crystals (diffraction limit varying from 2.90 to 3.25 Å) and from multiple orientations of the same crystal, using the κ-geometry goniostat on SOLEIL beamline PROXIMA 1, to obtain `true redundancy'. Data from three different crystals were combined for structure determination. An optimized HK construct bearing a shorter cloning artifact yielded crystals that diffracted X-rays to 2.51 Å resolution and that were used for final refinement of the model. Moreover, a thorougha posteriorianalysis using several different combinations of data sets allowed us to investigate the impact of the data-collection strategy on the success of the structure determination.

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Integrative structural Investigation on the architecture of human Importin4_histone H3/H4_Asf1a complex and its histone H3 tail binding

10.1101/181321 ◽

2017 ◽

Author(s):

Jungmin Yoon ◽

Seung Joong Kim ◽

Sojin An ◽

Alexander Leitner ◽

Taeyang Jung ◽

...

Keyword(s):

Histone H3 ◽

Terminal Region ◽

Nuclear Localization Sequence ◽

Molecular Architecture ◽

X Ray ◽

X Ray Crystallography ◽

Integrative Modeling ◽

Negative Stain ◽

Localization Sequence ◽

Chaperone Complex

AbstractImportin4 transports histone H3/H4 in complex with Asf1a to the nucleus for chromatin assembly. Importin4 recognizes the nuclear localization sequence located at the N-terminal tail of histones. Here, we analyzed the structures and interactions of human Importin4, histones and Asf1a by cross-linking mass spectrometry, X-ray crystallography, negative-stain electron microscopy, small-angle X-ray scattering and integrative modeling. The XL-MS data showed that the C-terminal region of Importin4 interacts extensively with the histone H3 tail. We determined the crystal structure of the C-terminal region of Importin4 bound to the histone H3 peptide, thus revealing that the acidic path in Importin4 accommodates the histone H3 tail and that histone H3 Lys14 is the primary residue interacting with Importin4. Furthermore, the molecular architecture of the Importin4_histone H3/H4_Asf1a complex was produced through an integrative modeling approach. Overall, this work provides structural insights into how Importin4 recognizes histones and their chaperone complex.

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Molecular architecture of the yeast Mediator complex

eLife ◽

10.7554/elife.08719 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 95

Author(s):

Philip J Robinson ◽

Michael J Trnka ◽

Riccardo Pellarin ◽

Charles H Greenberg ◽

David A Bushnell ◽

...

Keyword(s):

Electron Microscopy ◽

Structural Information ◽

Internal Controls ◽

Mediator Complex ◽

Amino Acid Residues ◽

Molecular Architecture ◽

Structural Elements ◽

X Ray ◽

X Ray Crystallography ◽

D Model

The 21-subunit Mediator complex transduces regulatory information from enhancers to promoters, and performs an essential role in the initiation of transcription in all eukaryotes. Structural information on two-thirds of the complex has been limited to coarse subunit mapping onto 2-D images from electron micrographs. We have performed chemical cross-linking and mass spectrometry, and combined the results with information from X-ray crystallography, homology modeling, and cryo-electron microscopy by an integrative modeling approach to determine a 3-D model of the entire Mediator complex. The approach is validated by the use of X-ray crystal structures as internal controls and by consistency with previous results from electron microscopy and yeast two-hybrid screens. The model shows the locations and orientations of all Mediator subunits, as well as subunit interfaces and some secondary structural elements. Segments of 20–40 amino acid residues are placed with an average precision of 20 Å. The model reveals roles of individual subunits in the organization of the complex.

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From Penicillin to the Ribosome: Revolutions in the Determination and Use of Molecular Structure in Chemistry and Biology

Australian Journal of Chemistry ◽

10.1071/ch04090 ◽

2004 ◽

Vol 57 (9) ◽

pp. 829 ◽

Cited By ~ 2

Author(s):

Edward N. Baker

Keyword(s):

Biological Sciences ◽

Molecular Structure ◽

Structural Genomics ◽

Structure Determination ◽

Biological Function ◽

Structural Information ◽

Structure Based Drug Design ◽

X Ray ◽

X Ray Crystallography

A revolution in structural analysis is in progress in the biological sciences that parallels a similar revolution that took place in chemistry 40–50 years ago. This has major implications for chemistry, offering exciting opportunities at the interface between chemistry and biology. The advances are driven by the value of structural information in biology, for understanding biological function, and for applications in structure-based drug design and structural genomics. Two directions are apparent: towards technically challenging biological structures and assemblies, typified by the potassium channel and the ribosome; and towards high-throughput structure determination of many, smaller, proteins, as in structural genomics. In this review, the advances in molecular biology and in structure determination by X-ray crystallography that make these developments possible are discussed, together with appropriate examples.

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