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Protein aggregation is sequence specific, favoring self-assembly over cross-seeding with non-homologous sequences. Still, as the majority of proteins in a proteome are aggregation prone, the high level of homogeneity of protein inclusions in vivo both during recombinant overexpression and in disease remains surprising. To investigate the selectivity of protein aggregation in a proteomic context, we here compared the selectivity of aggregation-determined interactions with antibody binding. To that purpose, we synthesized biotin-labeled peptides, corresponding to aggregation-determining sequences of the bacterial protein β-galactosidase and two human disease biomarkers: C-reactive protein and prostate-specific antigen. We analyzed the selectivity of their interactions in Escherichia coli lysate, human serum and human seminal plasma, respectively, using a Western blot-like approach in which the aggregating peptides replace the conventional antibody. We observed specific peptide accumulation in the same bands detected by antibody staining. Combined spectroscopic and mutagenic studies confirmed accumulation resulted from binding of the peptide on the identical sequence of the immobilized target protein. Further, we analyzed the sequence redundancy of aggregating sequences and found that about 90% of them are unique within their proteome. As a result, the combined specificity and low sequence redundancy of aggregating sequences therefore contribute to the observed homogeneity of protein aggregation in vivo. This suggests that these intrinsic proteomic properties naturally compartmentalize aggregation events in sequence space. In the event of physiological stress, this might benefit the ability of cells to respond to proteostatic stress by allowing chaperones to focus on specific aggregation events rather than having to face systemic proteostatic failure., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Protein aggregation is geared by aggregation-prone regions that self-associate by β-strand interactions. Charged residues and prolines are enriched at the flanks of aggregation-prone regions resulting in decreased aggregation. It is still unclear what drives the overrepresentation of these "aggregation gatekeepers", that is, whether their presence results from structural constraints determining protein stability or whether they constitute a bona fide functional class selectively maintained to control protein aggregation. As functional residues are typically conserved regardless of their cost to protein stability, we compared sequence conservation and thermodynamic cost of these residues in 2659 protein families in Escherichia coli. Across protein families, we find gatekeepers to be under strong selective conservation while at the same time representing a significant thermodynamic cost to protein structure. This finding supports the notion that aggregation gatekeepers are not structurally determined but evolutionary selected to control protein aggregation., (Copyright © 2014 Elsevier Ltd. All rights reserved.)