Structural insights into loss of function of a pore forming toxin and its role in pneumococcal adaptation to an intracellular lifestyle

PLoS Pathog. 2020 Nov 20;16(11):e1009016. doi: 10.1371/journal.ppat.1009016. eCollection 2020 Nov.


The opportunistic pathogen Streptococcus pneumoniae has dual lifestyles: one of an asymptomatic colonizer in the human nasopharynx and the other of a deadly pathogen invading sterile host compartments. The latter triggers an overwhelming inflammatory response, partly driven via pore forming activity of the cholesterol dependent cytolysin (CDC), pneumolysin. Although pneumolysin-induced inflammation drives person-to-person transmission from nasopharynx, the primary reservoir for pneumococcus, it also contributes to high mortality rates, creating a bottleneck that hampers widespread bacterial dissemination, thus acting as a double-edged sword. Serotype 1 ST306, a widespread pneumococcal clone, harbours a non-hemolytic variant of pneumolysin (Ply-NH). Performing crystal structure analysis of Ply-NH, we identified Y150H and T172I as key substitutions responsible for loss of its pore forming activity. We uncovered a novel inter-molecular cation-π interaction, governing formation of the transmembrane β-hairpins (TMH) in the pore state of Ply, which can be extended to other CDCs. H150 in Ply-NH disrupts this interaction, while I172 provides structural rigidity to domain-3, through hydrophobic interactions, inhibiting TMH formation. Loss of pore forming activity enabled improved cellular invasion and autophagy evasion, promoting an atypical intracellular lifestyle for pneumococcus, a finding that was corroborated in in vivo infection models. Attenuation of inflammatory responses and tissue damage promoted tolerance of Ply-NH-expressing pneumococcus in the lower respiratory tract. Adoption of this altered lifestyle may be necessary for ST306 due to its limited nasopharyngeal carriage, with Ply-NH, aided partly by loss of its pore forming ability, facilitating a benign association of SPN in an alternative, intracellular host niche.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptation, Physiological*
  • Amino Acid Sequence
  • Animals
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Cell Membrane / microbiology
  • Cholesterol / metabolism
  • Cytoplasm / microbiology
  • Female
  • Humans
  • Inflammation / microbiology*
  • Loss of Function Mutation*
  • Mice
  • Models, Structural
  • Perforin / genetics
  • Perforin / metabolism
  • Pneumococcal Infections / microbiology*
  • Sequence Alignment
  • Streptococcus pneumoniae / genetics
  • Streptococcus pneumoniae / physiology*
  • Streptolysins / genetics
  • Streptolysins / metabolism*


  • Bacterial Proteins
  • Streptolysins
  • plY protein, Streptococcus pneumoniae
  • Perforin
  • Cholesterol