Zacharchenko, T, Barsukov, I, Rigden, DJ et al. (2 more authors) (2016) Biophysical Analysis of the N-Terminal Domain from the Human Protein Phosphatase 1 Nuclear Targeting Subunit PNUTS Suggests an Extended Transcription Factor TFIIS-Like Fold. Protein Journal, 35 (5). pp. 340-345. ISSN 1572-3887
Abstract
Human protein phosphatase 1 nuclear targeting subunit (PNUTS) plays critical roles in DNA repair, cell growth and survival. The N-terminal domain of PNUTS mediates interactions with Tox4 and the phosphatase and tensin homolog PTEN, which are essential for the roles of this protein. To study this N-terminal domain, we have established its recombinant overproduction in E. coli utilizing NusA fusion. Upon removal of the tag, the remaining PNUTS sample is soluble and highly pure. We have characterized the domain using circular dichroism and nuclear magnetic resonance and analyzed its sequence using bioinformatics. All data agree in suggesting that the PNUTS N-terminal segment adopts a compact, globular fold rich in α-helical content, where the folded fraction is substantially larger than the previously annotated fold. We conclude that this domain adopts a single fold, likely being an extended form of the transcription factor S-II leucine/tryptophan conserved-motif. Thermal denaturation yielded a melting temperature of ~49.5 °C, confirming the stability of the fold. These findings pave the way for the molecular characterization of functional interactions mediated by the N-terminal region of PNUTS.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2016, Springer Science+Business Media New York. This is an author produced version of a paper published in Protein Journal. The final version is available at Springer via https://doi.org/10.1007/s10930-016-9677-7. Uploaded in accordance with the publisher's self-archiving policy. |
Keywords: | Recombinant protein overexpression; Secondary structure prediction; Thermal denaturation; Circular dichroism; Nuclear magnetic resonance |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Biology (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 28 Nov 2016 12:15 |
Last Modified: | 07 Sep 2017 07:28 |
Published Version: | https://doi.org/10.1007/s10930-016-9677-7 |
Status: | Published |
Publisher: | Springer Verlag |
Identification Number: | 10.1007/s10930-016-9677-7 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:108536 |