Sequence characteristics responsible for protein‐protein interactions in the intrinsically disordered regions of caseins, amelogenins, and small heat‐shock proteins

Abstract

Milk caseins and dental amelogenins are intrinsically disordered proteins (IDPs) that associate with themselves and others. Paradoxically, they are also described as hydrophobic proteins, which is difficult to reconcile with a solvent‐exposed conformation. We attempt to resolve this paradox. We show that caseins and amelogenins are not hydrophobic proteins but they are more hydrophobic than most IDPs. Remarkably, uncharged residues from different regions of these mature proteins have a nearly constant average hydropathy but these regions exhibit different charged residue frequencies. A novel sequence analysis method was developed to identify hydrophobic and order‐promoting regions that would favor conformational collapse. We found that such regions were uncommon; most hydrophobic and order‐promoting residues were adjacent to hydrophilic or disorder‐promoting residues. A further reason why caseins and amelogenins do not collapse is their high proportion of disorder‐promoting proline residues. We conclude that in these proteins the hydrophobic effect is not large enough to cause conformational collapse but it can contribute, along with polar interactions, to protein‐protein interactions. This behaviour is similar to the interaction of the disordered N‐terminal region of small heat‐shock proteins with either themselves during oligomer formation or other, unfolding, proteins during chaperone action.

The molecular chaperone β-casein prevents amorphous and fibrillar aggregation of α-lactalbumin by stabilisation of dynamic disorder

“…In general, molecular chaperones are classified into two classes, namely stabilising 'holdases' [40] and energy dependant 'foldases'. In agreement with previous studies [21][22][23][24]41,42], our data are consistent with β-CN functioning as a holdase to prevent protein aggregation without refolding aggregation-prone proteins to their native state.…”

“…Notably, this occurs without HMW complexation and despite the reduced hydrophobicity of RCM α-LA ( Figure 2). In vivo, β-CN associated with the other caseins (of which there are three in bovine milk) to form the large amorphous aggregate known as the casein micelle [22][23][24]. Consistent with their intrinsically disordered nature, β-CN and the other caseins adopt a significant proportion of polyproline-II (PP-II) helical secondary structure, mostly within their proline, glutamine-rich regions [22][23][24].…”

“…In vivo, β-CN associated with the other caseins (of which there are three in bovine milk) to form the large amorphous aggregate known as the casein micelle [22][23][24]. Consistent with their intrinsically disordered nature, β-CN and the other caseins adopt a significant proportion of polyproline-II (PP-II) helical secondary structure, mostly within their proline, glutamine-rich regions [22][23][24]. These regions are involved in casein-casein interactions within the casein micelle and with other proteins during chaperone action [22][23][24].…”

“…Like the other bovine caseins, β-CN exhibits chaperone activity to prevent the unfolding and amorphous aggregation of proteins under stress conditions such as elevated temperature [21]. β-CN also prevents amyloid fibril formation of κ-casein [22][23][24].…”

Dysregulated interactions triggered by a neuropathy-causing mutation in the IPV motif of HSP27

“…In addition, it has been demonstrated that other regions of sHSPs can also interact with the ACD, including NTD-β4/β8 groove interactions observed in a crystal structure of HSPB6 (69) and NTD-ACD dimer interface contacts as seen in a crystal structure of the HSPB2-HSPB3 hetero-tetramer (70). The CTRs of sHSPs play key roles in promoting solubility and contributing to the regulation of interactions with other proteins (71,72), similar to the roles of other intrinsically disordered regions (73). In addition, the CTR of HSPB2 was shown to regulate its ability to undergo liquid-liquid phase separation (74).…”