Relating Collapse to Sequence and Behavior in Glutamine‐Rich Domains

Abstract

Intrinsically disordered proteins, or IDPs, are ubiquitous throughout eukaryotic proteomes with particularly important roles in biosignalling and transcription regulation. Within this category, glutamine‐rich (Q‐rich) domains appear commonly within transcription activators. Q‐rich domains are known to collapse rather than fold upon binding and have a high tendency to aggregate. Their behavior appears to be mediated by solvent interaction with some evidence that water is trapped within the collapsed domains. These traits appear important in gene activation, yet effective pathways to evaluate the behavior of Q‐rich domains remain elusive. In this work, we develop fluorescence methods to experimentally observe the solvent exchange between bulk media and trapped water within collapsed Q‐rich domains. In conjunction with experimental observation, we have evaluated constructs for known parameters including disorder, solubility, and aggregation propensity to directly align experimental results with available prediction systems. We composed a library of disordered constructs from known transcription activators and repressors. Specifically, we selected peptides from within the Auxin response factor (ARF) family of proteins in Arabidopsis thaliana to utilize a biologically characterized genome with natural sequence variation including percent glutamine content. The control used was κ‐casein, an inexpensive protein that has been previously shown to collapse and trap water. We optimized a general expression, purification, and labeling protocol to isolate the library of constructs with a solvent‐sensitive fluorophore. Our methods allow us to directly observe both solvent exposure and exchange on constructs of varying glutamine content. This work provides important new tools to measure and compare Q‐rich domains. These tools can be applied to clarify the relationship between sequence and behavior in Q‐rich domains, specifically the biophysical implications of peptide‐solvent interactions.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.