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Updated: Jun 26, 2025
Quantifying the Cytotoxicity of Staphylococcus aureus Against Human Polymorphonuclear Leukocytes
Published on: January 3, 2020
Cytotoxic Staphylococcus aureus PSMα3 inhibits the aggregation of human insulin in vitro
Aleksandra Kalitnik1, Monika Szefczyk2, Alicja W Wojciechowska1
1Department of Biomedical Engineering, Faculty of Fundamental Problems of Technology, Wroclaw University of Science and Technology, Wybrzeże Wyspiańskiego 27, 50-370 Wroclaw, Poland. aleksandra.kalitnik@pwr.edu.pl.
Abstract:
Phenol-soluble modulins (PSMs) are extracellular short amphipathic peptides secreted by the bacteria Staphylococcus aureus (S. aureus). They play an essential role in the bacterial lifecycle, biofilm formation, and stabilisation. From the PSM family, PSMα3 has been of special interest recently due to its cytotoxicity and highly stable α-helical conformation, which also remains in its amyloid fibrils. In particular, PSMα3 fibrils were shown to be composed of self-associating "sheets" of α-helices oriented perpendicular to the fibril axis, mimicking the architecture of canonical cross-β fibrils. Therefore, they were called cross-α-fibrils. PSMα3 was synthesised and verified for identity with wild-type sequences (S. aureus). Then, using several experimental techniques, we evaluated its propensity for in vitro aggregation. According to our findings, synthetic PSMα3 (which lacks the N-terminal formyl groups found in bacteria) does not form amyloid fibrils and maintains α-helical conformation in a soluble monomeric form for several days of incubation. We also evaluated the influence of PSMα3 on human insulin fibrillation in vitro, using a variety of experimental approaches in combination with computational molecular studies. First, it was shown that PSMα3 drastically inhibits the fibrillation of human insulin. The anti-fibrillation effect of PSMα3 was concentration-dependent and required a concentration ratio of PSMα3: insulin equal to or above 1 : 100. Molecular modelling revealed that PSMα3 most likely inhibits the production of insulin primary nuclei by competing for residues involved in its dimerization.
Insights
Phenol-soluble modulin alpha 3 (PSMα3) from Staphylococcus aureus does not form amyloid fibrils on its own. However, it effectively inhibits human insulin fibrillation in a concentration-dependent manner.
Area of Science:
- Biochemistry
- Microbiology
- Structural Biology
Background:
- Phenol-soluble modulins (PSMs) are amphipathic peptides secreted by *Staphylococcus aureus*.
- PSMs are crucial for bacterial biofilm formation and stability.
- PSMα3 exhibits a stable α-helical conformation, even within amyloid fibrils, termed cross-α-fibrils.
Purpose of the Study:
- To investigate the in vitro aggregation propensity of synthetic PSMα3.
- To evaluate the effect of PSMα3 on human insulin fibrillation.
- To elucidate the mechanism by which PSMα3 influences insulin aggregation.
Main Methods:
- Synthesis and verification of wild-type PSMα3 sequence.
- In vitro aggregation assays using various experimental techniques.
- Computational molecular modeling studies.
- Insulin fibrillation assays with varying PSMα3 concentrations.
Main Results:
- Synthetic PSMα3, lacking N-terminal formyl groups, remained soluble and α-helical in vitro.
- PSMα3 significantly inhibited human insulin fibrillation in a concentration-dependent manner.
- The anti-fibrillation effect was observed at PSMα3:insulin ratios of 1:100 or higher.
- Molecular modeling suggested PSMα3 inhibits insulin primary nuclei formation by competing for dimerization residues.
Conclusions:
- Synthetic PSMα3 does not self-assemble into amyloid fibrils under tested conditions.
- PSMα3 acts as a potent inhibitor of human insulin fibrillation.
- The inhibitory mechanism involves interference with the initial stages of insulin nucleus formation.
