Catarina Fernandes:
Role of charges in disordered protein complexes

Date: 01-12-2022    Supervisor: Birthe B. Kragelund



Biological systems are governed by charges and electrostatic interactions. Proteins that are usually involved in disease can many times be highly charged. S100A13 is a Ca2+-binding protein that is involved in many important biological processes and has a characteristic highly positively charged patch on one side of its surface. Recently, it has been suggested that S00A13 plays a role as a transport chaperone of another highly charged protein, ProTα, which is intrinsically disordered, carrying an overall charge of -43. Once exported, ProTα has been suggested to exert a neuroprotection effect upon ischemia. However, how the two proteins engage in complex and how charges contribute to the binding mechanism remains to be known.

To begin to understand these types of electrostatic complexes we first described the context for protein interactions. Here, we highlight the importance of SLiMS and flanking regions for binding of IDPs as well as the context of the environment which includes pH, temperature and charges. Then, we asked if highly negatively charged IDPs have Ca2+ binding capabilities. With an increasingly number of negatively charged IDPs, there is a need to understand if particular binding motifs for Ca2+ were present. We showed that negatively charged IDPs bind Ca2+ using at least two different motifs binding the metal ion dynamically. 

Furthermore, we also investigated the role of charges for the stability and structure of the positively charged globular domain of histone H1.0. Here, we found that the addition or removal of charges did not disturb the structure of the protein but impacted its stability. Finally, using a combination of NMR, SAXS and smFRET we studied the interaction of S100A13 and ProTα. Our results suggest that ProTα binds S100A13 through its most negatively charged region at the interface of the S100A13 dimer localizing to the highly positively charged surface with no persistent contacts between specific residues. Thus, ProTα remains disordered but do so over restricted areas of the S100A13 surface. The binding region on S100A13 has not previously been shown to engage in ligand binding.

The work presented in this thesis provides important new insight into the role of charges in several different protein complexes representing different biological systems. The involvement of a high number of charges tend to enable more dynamic complexes. This provides certain biological advantages such as access for invading ligands and ligand exchange, access for modifying enzymes and dynamical scavenging of metal ions.