Aggregation gatekeepers modulate protein homeostasis of aggregating sequences and affect bacterial fitness

Protein Eng Des Sel. 2012 Jul;25(7):357-66. doi: 10.1093/protein/gzs031. Epub 2012 Jun 15.


The most common mechanism by which proteins aggregate consists in the assembly of short hydrophobic primary sequence segments into extended β-structured agglomerates. A significant enrichment of charged residues is observed at the flank of these aggregation-prone sequence segments, suggesting selective pressure against aggregation. These so-called aggregation gatekeepers act by increasing the intrinsic solubility of aggregating sequences in vitro, but it has been suggested that they could also facilitate chaperone interactions. Here, we address whether aggregation gatekeepers affect bacterial fitness. In Escherichia coli MC4100 we overexpressed GFP fusions with an aggregation-prone segment of σ32 (further termed σ32β) flanked by gatekeeper and non-gatekeeper residues and measured pairwise competitive growth. We found that the identity of flanking residues had significant effect on bacterial growth. Overexpression of σ32β flanked by its natural gatekeepers displayed the greatest competitive fitness, followed by other combinations of gatekeepers, while absence of gatekeepers strongly affects bacterial fitness. Further analysis showed the diversity of effects of gatekeepers on the proteostasis of σ32β including synthesis and degradation rates, in vivo aggregation propensity and chaperone response. Our results suggest that gatekeeper residues affect bacterial fitness not only by modulating the intrinsic aggregation propensity of proteins but also by the manner in which they affect the processing of σ32β-GFP by the protein quality control machinery of the cell. In view of these observations, we hypothesize that variation at gatekeeper positions offers a flexible selective strategy to modulate the proteostatic regulation of proteins to the match intrinsic aggregation propensities of proteins with required expression levels.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Escherichia coli / genetics
  • Escherichia coli / growth & development*
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism*
  • Gene Expression
  • Green Fluorescent Proteins / chemistry
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism*
  • Heat-Shock Proteins / chemistry
  • Heat-Shock Proteins / genetics
  • Heat-Shock Proteins / metabolism*
  • Hydrophobic and Hydrophilic Interactions
  • Protein Folding
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Sigma Factor / chemistry
  • Sigma Factor / genetics
  • Sigma Factor / metabolism*
  • Solubility
  • Up-Regulation


  • Escherichia coli Proteins
  • Heat-Shock Proteins
  • Recombinant Fusion Proteins
  • Sigma Factor
  • heat-shock sigma factor 32
  • Green Fluorescent Proteins