Bischofberger, Mirko; Iacovache, Mircea Ioan; Boss, Daniel; Naef, Felix; van der Goot, F Gisou; Molina, Nacho (2016). Revealing Assembly of a Pore-Forming Complex Using Single-Cell Kinetic Analysis and Modeling. Biophysical journal, 110(7), pp. 1574-1581. Biophysical Society 10.1016/j.bpj.2016.02.035
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Many biological processes depend on the sequential assembly of protein complexes. However, studying the kinetics of such processes by direct methods is often not feasible. As an important class of such protein complexes, pore-forming toxins start their journey as soluble monomeric proteins, and oligomerize into transmembrane complexes to eventually form pores in the target cell membrane. Here, we monitored pore formation kinetics for the well-characterized bacterial pore-forming toxin aerolysin in single cells in real time to determine the lag times leading to the formation of the first functional pores per cell. Probabilistic modeling of these lag times revealed that one slow and seven equally fast rate-limiting reactions best explain the overall pore formation kinetics. The model predicted that monomer activation is the rate-limiting step for the entire pore formation process. We hypothesized that this could be through release of a propeptide and indeed found that peptide removal abolished these steps. This study illustrates how stochasticity in the kinetics of a complex process can be exploited to identify rate-limiting mechanisms underlying multistep biomolecular assembly pathways.
Item Type: |
Journal Article (Original Article) |
|---|---|
Division/Institute: |
04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Anatomy |
UniBE Contributor: |
Iacovache, Mircea Ioan |
Subjects: |
500 Science > 570 Life sciences; biology 600 Technology > 610 Medicine & health |
ISSN: |
0006-3495 |
Publisher: |
Biophysical Society |
Language: |
English |
Submitter: |
Benoît Zuber |
Date Deposited: |
09 Jun 2016 14:18 |
Last Modified: |
05 Dec 2022 14:55 |
Publisher DOI: |
10.1016/j.bpj.2016.02.035 |
PubMed ID: |
27074682 |
BORIS DOI: |
10.7892/boris.81352 |
URI: |
https://boris.unibe.ch/id/eprint/81352 |
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