Intrinsically disordered stress proteins have been shown to act as chaperones, protecting proteins from damage caused by stresses such as freezing and thawing. Dehydration proteins (dehydrins) are intrinsically disordered stress proteins that are found in almost all land plants. They consist of a variable number of the short, semi-conserved, Y-, S-, and K-segments, with longer stretches of poorly conserved sequences in between. Previous studies have provided conflicting views on the details of the dehydrin cryoprotective mechanism of enzymes. Experiments with polyethylene glycol (PEG) have shown that PEG cryoprotective efficiency is the same as dehydrins of the same hydrodynamic radius, suggesting that the protein's disordered and polar nature is important, rather than the specific order of the residues. To further elucidate the mechanism, we created scrambled variants of the wild grape dehydrins K 2 and YSK 2 and tested their ability to protect lactate dehydrogenase and yeast frataxin homolog-1 from freeze/thaw damage. The results show that for preventing aggregation, it is the sequence composition and the size of the dehydrin that is the most important factor in protection, while for freeze/thaw damage causing loss of secondary structure, it is the sequence composition that is most significant.

Co-crystal structures of SPOP and Pdx1 peptides showed that Pdx1 occupies a canonical groove on SPOP, despite deviations in the Pdx1 sequence from other SPOP binding partners. Multivalency may also play a role in the binding affinity and, thus, regulation of Pdx1 stability by SPOP. Fluorescence titration experiments showed that, absent the other, the SLiMs within Pdx1C bind to SPOP with near millimolar dissociation constants; however, in the context of the full Pdx1 C-terminus, binding affinity increases dramatically. We demonstrate that, in cells, both SPOP binding sites within Pdx1 must be present to allow proteins to co-localize into nuclear speckles. An enhanced understanding of SPOP binding mechanisms with various substrates will yield valuable insight into the roles of SPOP and Pdx1 in etiologies of disease.