A "spindle and thread" mechanism unblocks p53 translation by modulating N-terminal disorder

Area of Science:

• Structural Biology
• Protein Biochemistry
• Cancer Therapeutics

Background:

• Disordered proteins, like tumor suppressor p53, present significant challenges in structural biology due to low expression and poor stability.
• These characteristics limit the development of effective cancer therapeutics targeting such proteins.
• p53's properties exemplify proteins existing precariously close to their solubility limits.

Purpose of the Study:

• To investigate if fusing p53 with a soluble spider silk domain (NT*) can modulate its expression and stability.
• To explore the potential of this fusion strategy for enhancing the translation and activity of disordered proteins.
• To understand the mechanism by which the spider silk domain impacts p53's biophysical properties.

Main Methods:

• Construction and expression of a chimeric protein fusing p53 with the NT* spider silk domain.
• Assessment of the chimeric protein's translation efficiency and functional activity in human cancer cells.
• Biophysical characterization to determine the conformation and interactions within the chimeric protein.

Main Results:

• The chimeric protein demonstrated highly efficient translation and retained full activity in cancer cells.
• Biophysical analysis revealed a compact structure with the disordered p53 transactivation domain wrapped around the NT* domain.
• The NT* domain was shown to facilitate the unblocking of p53's proline-rich disordered region during translation.

Conclusions:

• Fusion with the NT* spider silk domain enhances the co-translational folding and expression of disordered proteins.
• This 'spindle and thread' mechanism offers a novel strategy to overcome translation limitations for partially disordered proteins.
• The findings have implications for improving the therapeutic potential of disordered cancer targets.