PT  - JOURNAL ARTICLE
AU  - Michael J Senior
AU  - Carina Monico
AU  - Eve E Weatherill
AU  - Robert J Gilbert
AU  - Alejandro P Heuck
AU  - Mark I Wallace
TI  - Single-molecule imaging of cholesterol-dependent cytolysin assembly
AID  - 10.1101/2021.05.26.445776
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.05.26.445776
4099  - http://biorxiv.org/content/early/2021/05/27/2021.05.26.445776.short
4100  - http://biorxiv.org/content/early/2021/05/27/2021.05.26.445776.full
AB  - We exploit single-molecule tracking and optical single channel recording in droplet interface bilayers to resolve the assembly pathway of the Cholesterol-Dependent Cytolysin, Perfringolysin O. This enables quantification of the stoichiometry of PFO complexes during assembly with millisecond temporal resolution and 20 nanometre spatial precision. Our results support a model of overall stepwise irreversible assembly, dominated by monomer addition, but with infrequent assembly from larger partial complexes. Furthermore, our results suggest a dominant proportion of inserted, but non-conductive intermediates in assembly.Competing Interest StatementThe authors have declared no competing interest.