Abstract
Intrinsically disordered proteins (IDP) are proteins that sample a heterogeneous ensemble of conformers in solution. An estimated 25–30% of all eukaryotic proteins belong to this class. In vivo, IDPs function under conditions that are highly crowded by other biological macromolecules. Previous research has highlighted that the presence of crowding agents can influence the conformational ensemble sampled by IDPs, resulting in either compaction or expansion. The effects of self-crowding of the disordered protein Histatin 5 has, in an earlier study, been found to have limited influence on the conformational ensemble. In this study, it is examined whether the short chain length of Histatin 5 can explain the limited effects of crowding observed, by introducing (Histatin 5)2, a tandem repeat of Histatin 5. By utilizing small-angle X-ray scattering, it is shown that the conformational ensemble is conserved at high protein concentrations, in resemblance with Histatin 5, although with a lowered protein concentration at which aggregation arises. Under dilute conditions, atomistic molecular dynamics and coarse-grained Monte Carlo simulations, as well as an established scaling law, predicted more extended conformations than indicated by experimental data, hence implying that (Histatin 5)2 does not behave as a self-avoiding random walk.
Highlights
Disordered proteins (IDPs) lack a unique singular equilibrium structure; instead, they sample a heterogeneous ensemble of conformers in solution
Intrinsically disordered proteins (IDP) are often functional in environments that are highly crowded by other biological macromolecules, with cellular protein concentrations reaching as high as 400 mg/mL.[7]
In order to determine the effect of increased chain length on the propensity of secondary structure formation, bioinformatic analysis of the (Hst5)[2] sequence was performed by applying the PrDOS33 and IUPred2A32 algorithms
Summary
Disordered proteins (IDPs) lack a unique singular equilibrium structure; instead, they sample a heterogeneous ensemble of conformers in solution. IDPs retain a variety of biological functions[1] and have been estimated to account for 25−30% of all proteins in eukaryotic organisms.[2] Interactions of IDPs can be regulated by altering the affinity of the protein, through, for example, post-translational modifications, or by inducing changes to the conformational ensemble,[3,4] where the latter can be introduced by, for example, modifying the sequence length, the properties of the constituent amino acids, the presence of post-translational modifications, and the properties of the buffer such as ionic strength and pH.[5,6] In vivo, IDPs are often functional in environments that are highly crowded by other biological macromolecules, with cellular protein concentrations reaching as high as 400 mg/mL.[7]. An important factor observed is the excluded volume of both the crowding agent and the IDP.[17] Other factors that affect the crowding-induced effect observed include the linear charge density and the charge patterning of the IDP.[18]
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