Tryptophan (Trp) is the rarest of the 20 canonical amino acids; it is also the most metabolically costly to synthesize and the only amino acid other than methionine to be encoded by only one codon. These reasons have long been put forward to explain the low abundance of Trp in proteins. Here, we tested an alternative hypothesis, that Trp is used sparingly because it can readily promote polypeptide chain collapse and self-association, which can lead to aggregation. We were motivated to test this hypothesis in part due to the plethora of recent reports highlighting the important role of tyrosine (Tyr) residues in the formation of biomolecular condensates. The intrinsically disordered proteins (IDPs) and regions that lead to phase-separated, condensed states typically have low amino acid complexity and are enriched in Tyr residues, as well as other polar amino acids. In contrast, there has been relatively little focus on the contribution of Trp to phase separation and intermolecular interactions more generally, despite broad structural similarities between Trp and Tyr and general agreement that Trp is “stickier” than Tyr. To test the impact of Trp content and patterning on intra- and inter-molecular associations, we constructed a series of mutant sequences with different numbers and patterns of Trp residues, using a well-characterized intrinsically disordered protein as a template. We used a range of biophysical approaches to test the impact of these changes on intramolecular collapse and intermolecular associations. We found that the conformations of IDPs are sensitive to even subtle changes in Trp patterns. For example, the position of only a single Trp residue was sufficient to significantly stabilize intermolecular interactions. These results suggest the limited use of Trp is due to its adhesive properties, rather than metabolic cost.
Mukinay et al. (2026) studied this question.