Details
| Translated title of the contribution | Ein dimerer Holin/Antiholin-Komplex kontrolliert die Lyse durch den Phagen T4 |
|---|---|
| Original language | English |
| Article number | 1419106 |
| Journal | Frontiers in Microbiology |
| Volume | 15 |
| Publication status | Published - 5 Sept 2024 |
Abstract
release of cell wall-degrading endolysins. In phage T4, the antiholin RI inhibits
the holin T, thereby preventing the early release of the T4 endolysin and lysis.
The antiholin achieves lysis inhibition (LIN) in response to phage superinfections,
thereby increasing the chance for lysis in an environment with a lower phage
concentration. The holin T consists of a small N-terminal cytoplasmic domain,
a transmembrane helix, and a periplasmic C-terminal domain. The antiholin is
targeted to the periplasm by a cleavable signal peptide. Recently, the periplasmic
soluble domains of the holin and the antiholin were found to form T2/RI2
tetramers in crystals. To investigate the functional relevance of this complex,
we reconstituted LIN in a phage-free system, using only RI, T, and endolysin,
and combined targeted mutagenesis with functional analyses. Inactivation of
the RI signal peptide cleavage site did not abolish LIN, indicating that RI can
function in a membrane-bound state, which argued against the tetramer. This
led to analyses showing that only one of the two T/RI interfaces in the tetramer
is physiologically relevant, which is also the only interaction site predicted
by AlphaFold2. Some holin mutations at this interaction site prevented lysis,
suggesting that the RI interaction likely acts by blocking the holin oligomerization required for hole formation. We conclude that LIN is mediated by a dimeric T/RI complex that, unlike the tetramer, can be easily formed when both partners are membrane-anchored.
Keywords
- protein-protein interaction, membrane protein, bacteriophage, holin, antiholin, phage T4, lysis inhibition, Escherichia coli, protein–protein interaction
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- General Biochemistry,Genetics and Molecular Biology
- Medicine(all)
- Microbiology (medical)
- Immunology and Microbiology(all)
- Microbiology
Research Area (based on ÖFOS 2012)
- NATURAL SCIENCES
- Biology
- Biology
- Molecular biology
- NATURAL SCIENCES
- Biology
- Biology
- Microbiology
- NATURAL SCIENCES
- Biology
- Biology
- Structural biology
Cite this
- Standard
- Harvard
- Apa
- Vancouver
- BibTeX
- RIS
In: Frontiers in Microbiology, Vol. 15, 1419106, 05.09.2024.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - A dimeric holin/antiholin complex controls lysis by phage T4
AU - Schwarzkopf, Jan Michel Frederik
AU - Mehner-Breitfeld, Denise
AU - Brüser, Thomas
N1 - Publisher Copyright: Copyright © 2024 Schwarzkopf, Mehner-Breitfeld and Brüser.
PY - 2024/9/5
Y1 - 2024/9/5
N2 - Lytic phages control the timepoint of host cell lysis by timing the holin-mediatedrelease of cell wall-degrading endolysins. In phage T4, the antiholin RI inhibitsthe holin T, thereby preventing the early release of the T4 endolysin and lysis.The antiholin achieves lysis inhibition (LIN) in response to phage superinfections,thereby increasing the chance for lysis in an environment with a lower phageconcentration. The holin T consists of a small N-terminal cytoplasmic domain,a transmembrane helix, and a periplasmic C-terminal domain. The antiholin istargeted to the periplasm by a cleavable signal peptide. Recently, the periplasmicsoluble domains of the holin and the antiholin were found to form T2/RI2tetramers in crystals. To investigate the functional relevance of this complex,we reconstituted LIN in a phage-free system, using only RI, T, and endolysin,and combined targeted mutagenesis with functional analyses. Inactivation ofthe RI signal peptide cleavage site did not abolish LIN, indicating that RI canfunction in a membrane-bound state, which argued against the tetramer. Thisled to analyses showing that only one of the two T/RI interfaces in the tetrameris physiologically relevant, which is also the only interaction site predictedby AlphaFold2. Some holin mutations at this interaction site prevented lysis,suggesting that the RI interaction likely acts by blocking the holin oligomerization required for hole formation. We conclude that LIN is mediated by a dimeric T/RI complex that, unlike the tetramer, can be easily formed when both partners are membrane-anchored.
AB - Lytic phages control the timepoint of host cell lysis by timing the holin-mediatedrelease of cell wall-degrading endolysins. In phage T4, the antiholin RI inhibitsthe holin T, thereby preventing the early release of the T4 endolysin and lysis.The antiholin achieves lysis inhibition (LIN) in response to phage superinfections,thereby increasing the chance for lysis in an environment with a lower phageconcentration. The holin T consists of a small N-terminal cytoplasmic domain,a transmembrane helix, and a periplasmic C-terminal domain. The antiholin istargeted to the periplasm by a cleavable signal peptide. Recently, the periplasmicsoluble domains of the holin and the antiholin were found to form T2/RI2tetramers in crystals. To investigate the functional relevance of this complex,we reconstituted LIN in a phage-free system, using only RI, T, and endolysin,and combined targeted mutagenesis with functional analyses. Inactivation ofthe RI signal peptide cleavage site did not abolish LIN, indicating that RI canfunction in a membrane-bound state, which argued against the tetramer. Thisled to analyses showing that only one of the two T/RI interfaces in the tetrameris physiologically relevant, which is also the only interaction site predictedby AlphaFold2. Some holin mutations at this interaction site prevented lysis,suggesting that the RI interaction likely acts by blocking the holin oligomerization required for hole formation. We conclude that LIN is mediated by a dimeric T/RI complex that, unlike the tetramer, can be easily formed when both partners are membrane-anchored.
KW - Protein-Protein Interaktionen
KW - Membranproteine
KW - Bacteriophagen
KW - Holine
KW - Antiholine
KW - Phage T4
KW - Lysisinhibition
KW - Escherichia coli
KW - protein-protein interaction
KW - membrane protein
KW - bacteriophage
KW - holin
KW - antiholin
KW - phage T4
KW - lysis inhibition
KW - Escherichia coli
KW - protein–protein interaction
UR - http://www.scopus.com/inward/record.url?scp=85204743971&partnerID=8YFLogxK
U2 - 10.3389/fmicb.2024.1419106
DO - 10.3389/fmicb.2024.1419106
M3 - Article
VL - 15
JO - Frontiers in Microbiology
JF - Frontiers in Microbiology
SN - 1664-302X
M1 - 1419106
ER -