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Published on: May 13, 2020
Crystallization and preliminary X-ray diffraction analysis of the dimerization domain of the tumour suppressor ING4
Simone Culurgioni1, Inés G Muñoz, Alicia Palacios
1Structural Biology Unit, CIC bioGUNE, Parque Tecnológico de Bizkaia, Edificio 800, 48160 Derio, Spain.
Abstract:
Inhibitor of growth protein 4 (ING4) belongs to the ING family of tumour suppressors and is involved in chromatin remodelling, in growth arrest and, in cooperation with p53, in senescence and apoptosis. Whereas the structure and histone H3-binding properties of the C-terminal PHD domains of the ING proteins are known, no structural information is available for the N-terminal domains. This domain contains a putative oligomerization site rich in helical structure in the ING2-5 members of the family. The N-terminal domain of ING4 was overexpressed in Escherichia coli and purified to homogeneity. Crystallization experiments yielded crystals that were suitable for high-resolution X-ray diffraction analysis. The crystals belonged to the orthorhombic space group C222, with unit-cell parameters a = 129.7, b = 188.3, c = 62.7 A. The self-rotation function and the Matthews coefficient suggested the presence of three protein dimers per asymmetric unit. The crystals diffracted to a resolution of 2.3 A using synchrotron radiation at the Swiss Light Source (SLS) and the European Synchrotron Radiation Facility (ESRF).
Insights
The N-terminal domain of Inhibitor of Growth 4 (ING4) protein, a tumor suppressor, was structurally characterized. This research provides the first high-resolution X-ray diffraction data for this crucial domain.
Area of Science:
- Structural Biology
- Molecular Oncology
- Protein Crystallography
Background:
- Inhibitor of Growth 4 (ING4) is a tumor suppressor protein involved in chromatin remodeling, cell cycle arrest, senescence, and apoptosis.
- While the C-terminal domains of ING proteins are structurally characterized, the N-terminal domain, containing a putative helical oligomerization site, lacks structural information.
- Understanding ING4's N-terminal domain structure is crucial for elucidating its function in tumor suppression.
Purpose of the Study:
- To determine the high-resolution crystal structure of the N-terminal domain of Inhibitor of Growth 4 (ING4).
- To provide the first structural insights into the N-terminal domain of ING4.
- To facilitate further studies on ING4's oligomerization and tumor suppressor functions.
Main Methods:
- Overexpression and purification of the N-terminal domain of ING4 in Escherichia coli.
- Crystallization of the purified ING4 N-terminal domain.
- High-resolution X-ray diffraction analysis using synchrotron radiation at SLS and ESRF.
Main Results:
- Crystals of the ING4 N-terminal domain were obtained, belonging to the orthorhombic space group C222.
- The crystals diffracted to a resolution of 2.3 Å.
- Analysis suggested the presence of three protein dimers per asymmetric unit.
Conclusions:
- The study successfully obtained high-resolution X-ray diffraction data for the ING4 N-terminal domain.
- This structural information is a critical first step towards understanding the domain's role in ING4 function.
- The findings pave the way for future investigations into ING4's mechanism of action as a tumor suppressor.
