Membrane insertion mechanism and molecular assembly of the bacteriophage lysis toxin ΦX174-E.
Amino Acid Sequence
Antibiosis
/ genetics
Bacterial Proteins
/ genetics
Bacteriophage phi X 174
/ genetics
Binding Sites
Cell Wall
/ genetics
Dimyristoylphosphatidylcholine
/ chemistry
Escherichia coli
/ genetics
Escherichia coli Proteins
/ genetics
Gene Expression
Lipid Bilayers
/ chemistry
Lysogeny
/ genetics
Nanoparticles
/ chemistry
Peptidylprolyl Isomerase
/ genetics
Phosphatidylglycerols
/ chemistry
Protein Binding
Protein Conformation
Protein Engineering
/ methods
Protein Interaction Domains and Motifs
Protein Multimerization
Recombinant Proteins
/ chemistry
Sequence Alignment
Sequence Homology, Amino Acid
Solubility
Toxins, Biological
/ chemistry
Transferases (Other Substituted Phosphate Groups)
/ genetics
cell-free expression
molecular switch
nanodiscs
peptide antibiotics
phage lysis proteins
Journal
The FEBS journal
ISSN: 1742-4658
Titre abrégé: FEBS J
Pays: England
ID NLM: 101229646
Informations de publication
Date de publication:
05 2021
05 2021
Historique:
revised:
23
10
2020
received:
17
08
2020
accepted:
02
11
2020
pubmed:
28
11
2020
medline:
22
7
2021
entrez:
27
11
2020
Statut:
ppublish
Résumé
The bacteriophage ΦX174 causes large pore formation in Escherichia coli and related bacteria. Lysis is mediated by the small membrane-bound toxin ΦX174-E, which is composed of a transmembrane domain and a soluble domain. The toxin requires activation by the bacterial chaperone SlyD and inhibits the cell wall precursor forming enzyme MraY. Bacterial cell wall biosynthesis is an important target for antibiotics; therefore, knowledge of molecular details in the ΦX174-E lysis pathway could help to identify new mechanisms and sites of action. In this study, cell-free expression and nanoparticle technology were combined to avoid toxic effects upon ΦX174-E synthesis, resulting in the efficient production of a functional full-length toxin and engineered derivatives. Pre-assembled nanodiscs were used to study ΦX174-E function in defined lipid environments and to analyze its membrane insertion mechanisms. The conformation of the soluble domain of ΦX174-E was identified as a central trigger for membrane insertion, as well as for the oligomeric assembly of the toxin. Stable complex formation of the soluble domain with SlyD is essential to keep nascent ΦX174-E in a conformation competent for membrane insertion. Once inserted into the membrane, ΦX174-E assembles into high-order complexes via its transmembrane domain and oligomerization depends on the presence of an essential proline residue at position 21. The data presented here support a model where an initial contact of the nascent ΦX174-E transmembrane domain with the peptidyl-prolyl isomerase domain of SlyD is essential to allow a subsequent stable interaction of SlyD with the ΦX174-E soluble domain for the generation of a membrane insertion competent toxin.
Substances chimiques
Bacterial Proteins
0
Escherichia coli Proteins
0
Lipid Bilayers
0
Phosphatidylglycerols
0
Recombinant Proteins
0
SlyD protein, E coli
0
Toxins, Biological
0
1,2-dioleoyl-sn-glycero-3-phosphoglycerol
66322-31-4
1-palmitoyl-2-oleoylglycero-3-phosphoglycerol
81490-05-3
dimyristoylphosphatidylglycerol
BI71WT9P3R
Transferases (Other Substituted Phosphate Groups)
EC 2.7.8.-
mraY protein, Bacteria
EC 2.7.8.13
Peptidylprolyl Isomerase
EC 5.2.1.8
Dimyristoylphosphatidylcholine
U86ZGC74V5
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
3300-3316Informations de copyright
© 2020 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.
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