T4 endolysin (gene E product, also known as T4 lysozyme) is a GH24 family muramidase that catalyzes hydrolysis of the beta-1,4-glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in bacterial peptidoglycan. The enzyme functions in the periplasm after holin-mediated membrane permeabilization, degrading the peptidoglycan layer to enable viral release during the lytic cycle. It is one of the most extensively studied enzymes, with over 700 crystal structures deposited in the PDB. The catalytic mechanism involves a conserved Glu11-Asp20-Thr26 triad operating via an inverting glycosidase mechanism.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
|
GO:0003796
lysozyme activity
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: T4 endolysin has well-established lysozyme (muramidase) activity, catalyzing hydrolysis of the beta-1,4-glycosidic bond between N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG) in peptidoglycan [PMID:4865643]. The enzyme belongs to glycosyl hydrolase family 24 (GH24) with a conserved catalytic triad (Glu11-Asp20-Thr26). This IEA annotation is consistent with extensive experimental evidence.
Reason: Core molecular function of the enzyme. Lysozyme activity is definitively established by biochemical characterization [PMID:4865643] and structural studies showing substrate binding and catalytic mechanism [PMID:8266098]. The deep research review confirms T4 endolysin acts as an endo-acetylmuramidase cleaving glycosidic bonds at muramic acid residues.
Supporting Evidence:
PMID:1201752
T4 acts as an endo-acetylmuramidase capable of cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains.
|
|
GO:0003824
catalytic activity
|
IEA
GO_REF:0000043 |
ACCEPT |
Summary: General parent term for catalytic activity. T4 lysozyme is unequivocally a catalyst, but this term is very general and adds little information beyond the more specific lysozyme activity annotation.
Reason: While redundant with more specific terms, this IEA annotation from keyword mapping is not incorrect. It serves as a catch-all parent term. The more informative GO:0003796 (lysozyme activity) provides the specific function.
|
|
GO:0009253
peptidoglycan catabolic process
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: T4 lysozyme degrades peptidoglycan by hydrolyzing glycosidic bonds in the polysaccharide backbone. This catabolic activity is central to its biological role in enabling phage release by degrading the host cell wall.
Reason: Core biological process annotation. The enzyme directly catalyzes peptidoglycan degradation, which is its primary biological function. UniProt states it "degrades host peptidoglycans" [ECO:0000255|HAMAP-Rule:MF_04110]. The deep research confirms the enzyme "directly hydrolyzes peptidoglycan, weakening the sacculus."
Supporting Evidence:
PMID:1201752
Lysozyme from bacteriophage T4 was found to digest a soluble, uncrosslinked peptidoglycan
|
|
GO:0016787
hydrolase activity
|
IEA
GO_REF:0000043 |
ACCEPT |
Summary: General parent term for hydrolase activity. T4 lysozyme catalyzes hydrolysis of glycosidic bonds, so this parent term is correct but non-specific.
Reason: Correct parent term mapping. The enzyme is indeed a hydrolase (EC 3.2.1.17). While more specific terms are available (lysozyme activity, hydrolase activity acting on glycosyl bonds), this general IEA annotation is not wrong.
|
|
GO:0016798
hydrolase activity, acting on glycosyl bonds
|
IEA
GO_REF:0000043 |
ACCEPT |
Summary: T4 lysozyme hydrolyzes beta-1,4 glycosidic bonds between NAM and NAG residues. This term correctly captures the enzymatic mechanism at an intermediate level of specificity.
Reason: Appropriate parent term. T4 lysozyme cleaves glycosidic bonds as established by its EC number (3.2.1.17) and detailed structural/biochemical studies. This is more informative than generic hydrolase activity while being less specific than lysozyme activity.
|
|
GO:0016998
cell wall macromolecule catabolic process
|
IEA
GO_REF:0000002 |
ACCEPT |
Summary: Peptidoglycan is a major cell wall macromolecule in bacteria, and T4 lysozyme degrades it. This term is a parent of peptidoglycan catabolic process.
Reason: Correct parent term from InterPro mapping. The enzyme catabolizes peptidoglycan, which is the major structural component of bacterial cell walls. This is slightly less specific than GO:0009253 (peptidoglycan catabolic process) but correctly captures the biological process.
|
|
GO:0030430
host cell cytoplasm
|
IEA
GO_REF:0000044 |
ACCEPT |
Summary: T4 lysozyme accumulates in the host cell cytoplasm during viral replication before being released to the periplasm via holin-mediated membrane permeabilization.
Reason: Correct subcellular localization. UniProt states: "The endolysin is cytoplasmic, but can reach the periplasmic space with the help of the holins which disrupt the host cell membrane." The protein is synthesized and accumulates in the cytoplasm, making this a valid localization annotation.
Supporting Evidence:
file:BPT4/E/E-deep-research-falcon.md
T4 E endolysin is produced late, accumulates in the cytoplasm, and acts in the periplasm once holin T forms pores
|
|
GO:0031640
killing of cells of another organism
|
IEA
GO_REF:0000043 |
MARK AS OVER ANNOTATED |
Summary: This term describes "killing of cells of another organism" and was mapped from UniProt keywords. While T4 lysozyme activity does lead to host cell death, this framing is questionable for a phage-encoded enzyme that kills its own host cell as part of the viral life cycle.
Reason: This term is typically used for organisms with distinct identities where one kills cells of another (e.g., immune cells killing pathogens, antimicrobial peptides). For a phage enzyme, the host bacterium IS the organism in which the phage replicates - the relationship is more accurately described as "viral release from host cell" rather than killing another organism's cells. The term GO:0044659 (viral release from host cell by cytolysis) more accurately captures this biological context. This appears to be an over-annotation arising from keyword mapping.
|
|
GO:0042742
defense response to bacterium
|
IEA
GO_REF:0000043 |
REMOVE |
Summary: This term describes "reactions triggered in response to the presence of a bacterium that act to protect the cell or organism." This is fundamentally inappropriate for T4 lysozyme - a bacteriophage is not defending against bacteria; it is parasitizing them.
Reason: This is a clear case of inappropriate SPKW (SwissProt Keyword) over-annotation. The term GO:0042742 is designed for eukaryotic innate immunity and antimicrobial defense systems (e.g., lysozyme in tears defending against bacterial infection). T4 lysozyme is NOT a defense response - it is an offensive viral mechanism to lyse the host cell for progeny release. The phage does not "defend" against bacteria; it attacks and destroys them as part of its reproductive cycle. The correct process annotation is GO:0044659 (viral release from host cell by cytolysis). This annotation likely arose from the "Lysozyme" keyword being inappropriately linked to defense responses, without considering that phage lysozymes have the opposite biological context from eukaryotic lysozymes.
|
|
GO:0044659
viral release from host cell by cytolysis
|
IEA
GO_REF:0000104 |
ACCEPT |
Summary: T4 lysozyme is an essential component of the phage lysis system, working with holins and spanins to achieve programmed host cell lysis and release of mature virions.
Reason: Core biological process annotation. This accurately describes the primary biological role of T4 lysozyme. The deep research review states: "T4 E endolysin is produced late, accumulates in the cytoplasm, and acts in the periplasm once holin T forms pores; it directly hydrolyzes peptidoglycan, weakening the sacculus. Subsequent spanin action disrupts the outer membrane, completing lysis in Gram-negative hosts." UniProt confirms it "participates with the holin and spanin proteins in the sequential events which lead to the programmed host cell lysis releasing the mature viral particles."
Supporting Evidence:
file:BPT4/E/E-deep-research-falcon.md
Endolysin E functions in the periplasm following holin T pore formation; spanins mediate outer-membrane fusion/rupture to complete host-cell lysis in Gram-negative bacteria
|
|
GO:0003796
lysozyme activity
|
IDA
PMID:4865643 Purification of bacteriophage T4 lysozyme. |
ACCEPT |
Summary: Tsugita and Inouye (1968) purified T4 lysozyme and characterized its catalytic activity. This is the foundational experimental paper establishing the biochemical function.
Reason: Core experimental evidence for the primary molecular function. PMID:4865643 is cited in UniProt for the EC number assignment (EC=3.2.1.17) and catalytic activity characterization. This IDA annotation is well-supported by direct biochemical assays.
Supporting Evidence:
PMID:4865643
Purification of bacteriophage T4 lysozyme
|
|
GO:0003796
lysozyme activity
|
IDA
PMID:1201752 Studies on the specificity of action of bacteriophage T4 lys... |
ACCEPT |
Summary: Mirelman et al. (1975) characterized the substrate specificity of T4 lysozyme, demonstrating it acts as an endo-acetylmuramidase cleaving glycosidic bonds at peptide-substituted muramic acid residues.
Reason: Important experimental evidence providing mechanistic detail on substrate specificity. The paper directly demonstrates the muramidase activity and substrate requirements.
Supporting Evidence:
PMID:1201752
T4 acts as an endo-acetylmuramidase capable of cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains.
|
|
GO:0003796
lysozyme activity
|
IDA
PMID:4582731 Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine am... |
ACCEPT |
Summary: PMID:4582731 (Inouye et al. 1973) is titled "Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine amidase" and describes T7 lysozyme, NOT T4 lysozyme. This reference is incorrectly attributed to the T4 gene E product. The annotation itself (GO:0003796 lysozyme activity) is correct for T4 lysozyme based on other evidence, but this specific annotation line should be removed due to incorrect reference.
Reason: While this specific annotation line has an incorrect reference (PMID:4582731 is about T7 lysozyme, not T4 lysozyme), the GO term itself (GO:0003796 lysozyme activity) is unequivocally correct for T4 lysozyme based on extensive other evidence. The term should be ACCEPT because lysozyme activity is the core function of this enzyme. However, note that this specific reference attribution is erroneous - T4 and T7 lysozymes are unrelated enzymes with different catalytic mechanisms (T4 is a muramidase cleaving glycosidic bonds; T7 is an amidase cleaving peptide bonds). The annotation itself should remain, but the reference should be corrected in the source database.
Supporting Evidence:
PMID:4582731
Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine amidase
|
Q: Is there any evidence for substrate specificity differences between T4 lysozyme and other GH24 lysozymes?
Q: Are there documented cases of T4 lysozyme activity outside the lysis pathway context?
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template_variables:
organism: BPT4
gene_id: E
gene_symbol: E
uniprot_accession: P00720
protein_description: 'RecName: Full=Endolysin {ECO:0000255|HAMAP-Rule:MF_04110};
EC=3.2.1.17 {ECO:0000255|HAMAP-Rule:MF_04110, ECO:0000269|PubMed:4865643}; AltName:
Full=Lysis protein {ECO:0000255|HAMAP-Rule:MF_04110}; AltName: Full=Lysozyme {ECO:0000255|HAMAP-Rule:MF_04110};
AltName: Full=Muramidase {ECO:0000255|HAMAP-Rule:MF_04110};'
gene_info: Name=E;
organism_full: Enterobacteria phage T4 (Bacteriophage T4).
protein_family: Belongs to the glycosyl hydrolase 24 family.
protein_domains: Endolysin_T4_type. (IPR034690); Glyco_hydro_24. (IPR002196); Lysozyme-like_dom_sf.
(IPR023346); Lysozyme_dom_sf. (IPR023347); Phage_lysozyme-like. (IPR052619)
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citation_count: 24
BEFORE YOU BEGIN RESEARCH: You MUST verify you are researching the CORRECT gene/protein. Gene symbols can be ambiguous, especially for less well-characterized genes from non-model organisms.
DO NOT PROCEED WITH RESEARCH ON A DIFFERENT GENE. Instead:
- State clearly: "The gene symbol 'E' is ambiguous or literature is limited for this specific protein"
- Explain what you found (e.g., "Found extensive literature on a different gene with the same symbol in a different organism")
- Describe the protein based ONLY on the UniProt information provided above
- Suggest that the protein function can be inferred from domain/family information
Please provide a comprehensive research report on the gene E (gene ID: E, UniProt: P00720) in BPT4.
The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.
You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.
We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.
We are interested in where in or outside the cell the gene product carries out its function.
We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.
Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.
Question: You are an expert researcher providing comprehensive, well-cited information.
Provide detailed information focusing on:
1. Key concepts and definitions with current understanding
2. Recent developments and latest research (prioritize 2023-2024 sources)
3. Current applications and real-world implementations
4. Expert opinions and analysis from authoritative sources
5. Relevant statistics and data from recent studies
Format as a comprehensive research report with proper citations. Include URLs and publication dates where available.
Always prioritize recent, authoritative sources and provide specific citations for all major claims.
BEFORE YOU BEGIN RESEARCH: You MUST verify you are researching the CORRECT gene/protein. Gene symbols can be ambiguous, especially for less well-characterized genes from non-model organisms.
DO NOT PROCEED WITH RESEARCH ON A DIFFERENT GENE. Instead:
- State clearly: "The gene symbol 'E' is ambiguous or literature is limited for this specific protein"
- Explain what you found (e.g., "Found extensive literature on a different gene with the same symbol in a different organism")
- Describe the protein based ONLY on the UniProt information provided above
- Suggest that the protein function can be inferred from domain/family information
Please provide a comprehensive research report on the gene E (gene ID: E, UniProt: P00720) in BPT4.
The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.
You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.
We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.
We are interested in where in or outside the cell the gene product carries out its function.
We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.
Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.
Plan status: Identity verified and evidence gathered. Proceeding with a comprehensive, cited research report.
Gene/protein verification and scope
- Symbol and organism: The target is gene E of Enterobacteria phage T4 (Bacteriophage T4), encoding the canonical T4 endolysin, also known as T4 lysozyme (UniProt P00720). Literature consistently places T4 lysozyme in glycoside hydrolase family 24 (GH24) and the T4-type lysozyme-like superfamily; catalytic features and substrate specificity match this assignment (Jun 2021; Viruses) (love2021themolecularbasis pages 10-12, ramesh2022alysozymemurein pages 1-2). No conflicting gene symbol “E” usages from other organisms are used here.
Executive summary of essentials
| Aspect | Key findings | Recent/authoritative sources |
|---|---|---|
| Identity verification | Gene E (UniProt P00720) from Enterobacteria phage T4 encodes the canonical T4 endolysin / T4 lysozyme; assigned to glycosyl hydrolase family 24 (GH24) and T4-type lysozyme-like proteins. | (ramesh2022alysozymemurein pages 1-2, love2021themolecularbasis pages 10-12) |
| Catalytic activity | Muramidase activity: cleaves the β-1,4 glycosidic bond between N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG); conserved catalytic residues Glu11, Asp20, Thr26; T4L described as an inverting glycosidase (with T26H mutant converting to a retaining mechanism in structural studies). | (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14, schwarzkopf2025theshortcytoplasmic pages 1-2) |
| Structural family / domains | GH24 (lysozyme-like) fold with conserved motif (Glu–(X)8–Asp–(X)5–Thr) and the two-domain α/β architecture seen across T4L-like endolysins. | (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 18-19, ramesh2022alysozymemurein pages 1-2) |
| Localization / biological role | Endolysin acts on the peptidoglycan in the periplasm after holin-mediated translocation; spanins mediate outer-membrane disruption for explosive lysis in Gram-negative hosts. | (schwarzkopf2025theshortcytoplasmic pages 1-2, krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12) |
| Lysis regulation | Lysis timing controlled by holin T and antiholin RI; RI–T interactions mediate lysis inhibition (LIN), with DNA proposed as a LIN signal stabilizing RI–T complexes. | (krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12) |
| 2024–2025 advances | 2024: evidence for a physiologically relevant dimeric holin–antiholin (T/RI) complex controlling lysis; 2025: microfluidics/fluorescence and modeling indicate small holin assemblies (ring of dimers) suffice to form aqueous pores enabling endolysin release. | (schwarzkopf2024adimericholinantiholin pages 12-12, schwarzkopf2025theshortcytoplasmic pages 1-2) |
| Applications | Phage endolysins are being developed as antimicrobials; 2024 preclinical/animal efficacy example: LysSYL shows broad anti-staphylococcal activity and strong in vivo protection; clinical candidates (e.g., exebacase) progressed into late-stage studies (Phase 3 referenced). | (liu2024lyssylabroadspectrum pages 15-17, liu2024lyssylabroadspectrum pages 1-2, elst2026clinicalimplementationof pages 36-42) |
| Quantitative data | LysSYL: 6-log10 reduction in vitro at 50 µg/mL and 100% survival in mice at 50 mg/kg (single dose vs 1×10^8 CFU challenge); exebacase: Phase 3 clinical trial reported (see clinical pipeline citations). | (liu2024lyssylabroadspectrum pages 15-17, liu2024lyssylabroadspectrum pages 1-2, elst2026clinicalimplementationof pages 36-42) |
Table: A compact reference table summarizing identity, mechanism, structural class, localization and regulation, recent 2024–2025 advances, applications, and key quantitative results for the T4 endolysin (gene E, UniProt P00720), with provenance to cited sources.
1) Key concepts and definitions with current understanding
- Endolysin (phage lysin): Phage-encoded peptidoglycan hydrolase that degrades the bacterial cell wall at the end of the replication cycle to enable virion release. In Gram-negative hosts such as E. coli, endolysins act in the periplasm after holin-triggered access; outer-membrane rupture is mediated by spanins (2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
- T4 endolysin (gene E product; T4 lysozyme): A GH24 muramidase that cleaves the β-1,4-glycosidic bond between N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG) in peptidoglycan. Catalysis relies on a conserved triad Glu11–Asp20–Thr26, with an inverting mechanism in wild type; the T26H mutant exhibits a retaining mechanism in structural studies (Jun 2021; Viruses; 2017; Protein Science) (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
- Holin–endolysin–spanin lysis system: Holin T (T4) forms inner-membrane lesions at a programmed time, allowing endolysin E to reach the periplasmic peptidoglycan; spanins effect outer-membrane disruption for explosive lysis (2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
- Lysis inhibition (LIN): In T4, antiholin RI modulates holin T to delay lysis. Recent work proposes that DNA stabilizes the RI–T complex and acts as the LIN signal (Jul 2020; J Mol Biol) (krieger2020thestructuralbasis pages 1-3).
2) Primary function, catalytic activity, and structure of T4 endolysin (gene E)
- Reaction and substrate specificity: T4 E encodes a muramidase that hydrolyzes the β-1,4 NAM–NAG linkage in peptidoglycan, i.e., an N-acetyl-β-D-muramidase activity typical of GH24/T4-type lysozymes (Jun 2021; Viruses; 2025; Frontiers in Microbiology) (love2021themolecularbasis pages 10-12, schwarzkopf2025theshortcytoplasmic pages 1-2).
- Catalytic residues/mechanism: The T4L-like catalytic triad Glu11 (general acid), Asp20 (general base), and Thr26 (positions nucleophilic water) is conserved across T4L-like endolysins; T4L is characterized as an inverting glycosidase, while T26H yields a retaining mechanism in structural analysis (Jun 2021; Viruses; 2017; Protein Science) (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
- Structural family/domains: T4 endolysin belongs to GH24 and the lysozyme-like fold with a conserved motif Glu–(X)8–Asp–(X)5–Thr and a two-domain architecture (α-helix bundle plus catalytic domain); key positions that stabilize the catalytic glutamate and geometry are conserved across the family (Jun 2021; Viruses) (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
3) Localization and role in the lysis pathway (holin T, antiholin RI, spanins) and lysis inhibition
- Localization and timing: Endolysin E functions in the periplasm following holin T pore formation; spanins mediate outer-membrane fusion/rupture to complete host-cell lysis in Gram-negative bacteria (2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
- Holin pore formation: Single-cell microfluidics and fluorescence tracking indicate that small assemblies of holin T (rings of dimers) can form aqueous pores sufficient to release endolysin; large multimerization is not required for endolysin release, and the short N-terminal cytoplasmic region of holin T is essential for pore formation (May 2025; Frontiers in Microbiology; URL: https://doi.org/10.3389/fmicb.2025.1579756) (schwarzkopf2025theshortcytoplasmic pages 1-2).
- Lysis inhibition control: Structural and biochemical studies show RI–T complexes are stabilized by DNA, leading to the proposal that DNA (from superinfecting phage) acts as the LIN signal; RI also exhibits periplasmic targeting and regulated stability (Jul 2020; J Mol Biol; URL: https://doi.org/10.1016/j.jmb.2020.06.013) (krieger2020thestructuralbasis pages 1-3).
- Holin–antiholin stoichiometry in 2024: Functional analyses support a physiologically relevant dimeric T/RI complex for LIN, with membrane-anchored partners and mutations in the holin–RI interface abolishing lysis, arguing that RI blocks holin oligomerization required for pore formation (Sep 2024; Frontiers in Microbiology; URL: https://doi.org/10.3389/fmicb.2024.1419106) (schwarzkopf2024adimericholinantiholin pages 12-12).
4) Recent developments and latest research (prioritizing 2023–2024)
- Dimeric holin/antiholin complex in T4: 2024 experiments reconstituted LIN in a phage-free system (RI, T, endolysin) and, through mutagenesis and functional assays, concluded that a dimeric T/RI complex mediates LIN by preventing holin oligomerization (Sep 2024; Frontiers in Microbiology) (schwarzkopf2024adimericholinantiholin pages 12-12).
- DNA as the lysis-inhibition signal: 2020 structural work provided the mechanistic basis that DNA stabilizes the RI–T complex, reframing LIN signaling in T4 (Jul 2020; J Mol Biol) (krieger2020thestructuralbasis pages 1-3).
- Holin pore visualization and minimal assemblies: 2025 microfluidics and modeling show small holin assemblies suffice to form pores, rebalancing prior assumptions about large holin holes and emphasizing the essential role of holin’s short cytoplasmic region (May 2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
5) Current applications and real-world implementations
- Endolysin therapeutics: 2024 preclinical data describe LysSYL, an anti-staphylococcal endolysin, with broad in vitro activity (100% of tested Staphylococcus strains vs. parent phage’s 41.7%), rapid killing of MRSA USA300 within 10 minutes, biofilm eradication, anti-persister activity, and strong efficacy in a murine peritonitis model (Mar 2024; Microbial Cell Factories; URL: https://doi.org/10.1186/s12934-024-02359-4) (liu2024lyssylabroadspectrum pages 1-2, liu2024lyssylabroadspectrum pages 15-17).
- Clinical pipeline context: A 2026 selective review summarizes clinical development and combination strategies for Gram-positive endolysins, including the Phase 3 DISRUPT trial of exebacase reported in 2024 (Clinical Infectious Diseases, DOI listed therein), and multiple other clinical candidates (European Journal of Medical Research; URL: https://doi.org/10.1186/s40001-025-03655-4) (elst2026clinicalimplementationof pages 36-42, elst2026clinicalimplementationof pages 31-36).
- Relevance to T4 endolysin: T4 lysozyme is a paradigm GH24 enzyme and a common structural/biophysical model for engineering endolysins and understanding catalytic mechanisms used in antimicrobial design (Jun 2021; Viruses) (love2021themolecularbasis pages 10-12).
6) Expert opinions and analysis from authoritative sources
- Mechanistic paradigm: T4L remains a structural and mechanistic archetype for GH24 muramidases; conserved catalytic geometry and fold are re-used in many T4L-like endolysins, guiding rational engineering for stability and activity (Jun 2021; Viruses) (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
- Lysis control in T4: Structural biology (J Mol Biol 2020) and functional reconstitution (Frontiers 2024) converge on a model where DNA-stabilized RI–T complexes implement LIN by blocking holin oligomerization, refining decades-old models of lysis timing control (Jul 2020; J Mol Biol; Sep 2024; Frontiers in Microbiology) (krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12).
- Holin pore formation: Single-cell and modeling data support minimal pore assemblies for holin T, underscoring regulatory roles of holin domains and suggesting that endolysin availability influences holin multimerization states (May 2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
7) Relevant statistics and data from recent studies
- Catalytic mechanism and residues: The conserved T4L triad Glu–Asp–Thr is aligned in T4L-like endolysins; T4L is inverting, while T26H shows retaining features in structural studies (Jun 2021; Viruses; 2017; Protein Science) (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
- Lysis-regulation data: DNA-stabilized 2RI–2T complexes proposed for LIN (structures deposited); functional 2024 work supports a physiologically relevant dimeric T/RI complex under membrane-anchored conditions (Jul 2020; J Mol Biol; Sep 2024; Frontiers in Microbiology) (krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12).
- Therapeutic efficacy: LysSYL reduced MRSA by 6 log10 at 50 µg/mL in vitro and achieved 100% survival at 50 mg/kg in a mouse peritonitis model; LysSYL lysed 100% of 115 Staphylococcus strains vs. 41.7% for the parent phage (Mar 2024; Microbial Cell Factories) (liu2024lyssylabroadspectrum pages 15-17, liu2024lyssylabroadspectrum pages 1-2).
- Clinical status: Exebacase Phase 3 (2024; Clinical Infectious Diseases) and broader clinical landscape summarized with trial identifiers and combination strategy analysis (Jan 2026; European Journal of Medical Research) (elst2026clinicalimplementationof pages 36-42, elst2026clinicalimplementationof pages 31-36).
Biological pathway integration and localization (focused on precision of role)
- T4 E endolysin is produced late, accumulates in the cytoplasm, and acts in the periplasm once holin T forms pores; it directly hydrolyzes peptidoglycan, weakening the sacculus. Subsequent spanin action disrupts the outer membrane, completing lysis in Gram-negative hosts (2025; Frontiers in Microbiology) (schwarzkopf2025theshortcytoplasmic pages 1-2).
- Lysis timing is tuned by LIN: RI binds holin T; DNA stabilizes the complex to delay pore formation and endolysin release when superinfection occurs, thereby modulating the lytic schedule (Jul 2020; J Mol Biol; Sep 2024; Frontiers in Microbiology) (krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12).
Notes on applications and translational outlook
- As a well-characterized GH24 lysozyme, T4 endolysin remains a foundational model for designing and engineering T4L-like endolysins for therapeutic use, informing catalytic-site and stability modifications (Jun 2021; Viruses) (love2021themolecularbasis pages 10-12). 2024 preclinical data across the endolysin field (e.g., LysSYL) demonstrate rapid killing, biofilm clearance, and synergy with antibiotics in vivo; 2024 Phase 3 reporting for exebacase signals maturation of the field, with expert analyses suggesting combination strategies with standard-of-care antibiotics for systemic infections (Mar 2024; Microbial Cell Factories; Jan 2026; European Journal of Medical Research) (liu2024lyssylabroadspectrum pages 1-2, elst2026clinicalimplementationof pages 36-42, elst2026clinicalimplementationof pages 31-36).
URLs and publication dates (selection)
- DNA as LIN signal; J Mol Biol; published July 24, 2020: https://doi.org/10.1016/j.jmb.2020.06.013 (krieger2020thestructuralbasis pages 1-3).
- Dimeric T/RI complex controls lysis; Frontiers in Microbiology; Sep 2024: https://doi.org/10.3389/fmicb.2024.1419106 (schwarzkopf2024adimericholinantiholin pages 12-12).
- Holin pore formation by minimal assemblies; Frontiers in Microbiology; May 2025: https://doi.org/10.3389/fmicb.2025.1579756 (schwarzkopf2025theshortcytoplasmic pages 1-2).
- T4L-like catalytic mechanism/fold; Viruses; Jun 2021: https://doi.org/10.3390/v13061101 (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14).
- Endolysin efficacy (LysSYL); Microbial Cell Factories; Mar 2024: https://doi.org/10.1186/s12934-024-02359-4 (liu2024lyssylabroadspectrum pages 1-2, liu2024lyssylabroadspectrum pages 15-17).
- Clinical landscape and exebacase Phase 3 citation; European Journal of Medical Research; Jan 2026: https://doi.org/10.1186/s40001-025-03655-4 (elst2026clinicalimplementationof pages 36-42, elst2026clinicalimplementationof pages 31-36).
Conclusions
- The T4 gene E product is a GH24 muramidase that acts in the periplasm to cleave the NAM–NAG β-1,4 linkage, with conserved Glu11–Asp20–Thr26 catalysis and an inverting mechanism in wild type. Its role is tightly integrated into the holin–endolysin–spanin pathway and the LIN regulatory network, where recent work refined the molecular logic: DNA stabilization of RI–T underlies lysis inhibition, and small holin assemblies can form pores adequate for endolysin release. Translationally, T4L remains a benchmark mechanistic and structural template for engineering endolysins; 2023–2024 studies show compelling preclinical efficacy and advancing clinical programs, with expert analyses favoring combination strategies for systemic infections (love2021themolecularbasis pages 10-12, love2021themolecularbasis pages 12-14, schwarzkopf2025theshortcytoplasmic pages 1-2, krieger2020thestructuralbasis pages 1-3, schwarzkopf2024adimericholinantiholin pages 12-12, liu2024lyssylabroadspectrum pages 1-2, elst2026clinicalimplementationof pages 36-42, elst2026clinicalimplementationof pages 31-36).
References
(love2021themolecularbasis pages 10-12): Michael J. Love, David Coombes, Sarah H. Manners, Gayan S. Abeysekera, Craig Billington, and Renwick C. J. Dobson. The molecular basis for escherichia coli o157:h7 phage fahec1 endolysin function and protein engineering to increase thermal stability. Viruses, 13:1101, Jun 2021. URL: https://doi.org/10.3390/v13061101, doi:10.3390/v13061101. This article has 24 citations and is from a poor quality or predatory journal.
(ramesh2022alysozymemurein pages 1-2): Nachimuthu Ramesh, Prasanth Manohar, Kandasamy Eniyan, Loganathan Archana, Sudarsanan Athira, Belinda Loh, Long Ma, and Sebastian Leptihn. A lysozyme murein hydrolase with broad-spectrum antibacterial activity from enterobacter phage mypsh1140. Antimicrobial Agents and Chemotherapy, Sep 2022. URL: https://doi.org/10.1128/aac.00506-22, doi:10.1128/aac.00506-22. This article has 21 citations and is from a highest quality peer-reviewed journal.
(love2021themolecularbasis pages 12-14): Michael J. Love, David Coombes, Sarah H. Manners, Gayan S. Abeysekera, Craig Billington, and Renwick C. J. Dobson. The molecular basis for escherichia coli o157:h7 phage fahec1 endolysin function and protein engineering to increase thermal stability. Viruses, 13:1101, Jun 2021. URL: https://doi.org/10.3390/v13061101, doi:10.3390/v13061101. This article has 24 citations and is from a poor quality or predatory journal.
(schwarzkopf2025theshortcytoplasmic pages 1-2): Jan Michel Frederik Schwarzkopf, Ruth Paola Viveros, Ali Nazmi Burdur, Denise Mehner-Breitfeld, Natalia Tschowri, and Thomas Brüser. The short cytoplasmic region of phage t4 holin is essential for the transition from impermeable membrane protein complexes to permeable pores. Frontiers in Microbiology, May 2025. URL: https://doi.org/10.3389/fmicb.2025.1579756, doi:10.3389/fmicb.2025.1579756. This article has 1 citations and is from a poor quality or predatory journal.
(love2021themolecularbasis pages 18-19): Michael J. Love, David Coombes, Sarah H. Manners, Gayan S. Abeysekera, Craig Billington, and Renwick C. J. Dobson. The molecular basis for escherichia coli o157:h7 phage fahec1 endolysin function and protein engineering to increase thermal stability. Viruses, 13:1101, Jun 2021. URL: https://doi.org/10.3390/v13061101, doi:10.3390/v13061101. This article has 24 citations and is from a poor quality or predatory journal.
(krieger2020thestructuralbasis pages 1-3): Inna V. Krieger, Vladimir Kuznetsov, Jeng-Yih Chang, Junjie Zhang, Samir H. Moussa, Ryland F. Young, and James C. Sacchettini. The structural basis of t4 phage lysis control: dna as the signal for lysis inhibition. Journal of Molecular Biology, 432:4623-4636, Jul 2020. URL: https://doi.org/10.1016/j.jmb.2020.06.013, doi:10.1016/j.jmb.2020.06.013. This article has 35 citations and is from a domain leading peer-reviewed journal.
(schwarzkopf2024adimericholinantiholin pages 12-12): Jan Michel Frederik Schwarzkopf, Denise Mehner-Breitfeld, and Thomas Brüser. A dimeric holin/antiholin complex controls lysis by phage t4. Frontiers in Microbiology, Sep 2024. URL: https://doi.org/10.3389/fmicb.2024.1419106, doi:10.3389/fmicb.2024.1419106. This article has 10 citations and is from a poor quality or predatory journal.
(liu2024lyssylabroadspectrum pages 15-17): He Liu, Xuemei Wei, Zhefen Wang, Xiaonan Huang, Mengyang Li, Zhen Hu, Kexin Zhang, Qiwen Hu, Huagang Peng, Weilong Shang, Yi Yang, Yuting Wang, Shuguang Lu, and Xiancai Rao. Lyssyl: a broad-spectrum phage endolysin targeting staphylococcus species and eradicating s. aureus biofilms. Microbial Cell Factories, Mar 2024. URL: https://doi.org/10.1186/s12934-024-02359-4, doi:10.1186/s12934-024-02359-4. This article has 37 citations and is from a peer-reviewed journal.
(liu2024lyssylabroadspectrum pages 1-2): He Liu, Xuemei Wei, Zhefen Wang, Xiaonan Huang, Mengyang Li, Zhen Hu, Kexin Zhang, Qiwen Hu, Huagang Peng, Weilong Shang, Yi Yang, Yuting Wang, Shuguang Lu, and Xiancai Rao. Lyssyl: a broad-spectrum phage endolysin targeting staphylococcus species and eradicating s. aureus biofilms. Microbial Cell Factories, Mar 2024. URL: https://doi.org/10.1186/s12934-024-02359-4, doi:10.1186/s12934-024-02359-4. This article has 37 citations and is from a peer-reviewed journal.
(elst2026clinicalimplementationof pages 36-42): Niels Vander Elst. Clinical implementation of endolysins targeting gram-positive bacteria points toward a combination strategy with standard-of-care antibiotics: a selective review. European Journal of Medical Research, Jan 2026. URL: https://doi.org/10.1186/s40001-025-03655-4, doi:10.1186/s40001-025-03655-4. This article has 0 citations and is from a peer-reviewed journal.
(elst2026clinicalimplementationof pages 31-36): Niels Vander Elst. Clinical implementation of endolysins targeting gram-positive bacteria points toward a combination strategy with standard-of-care antibiotics: a selective review. European Journal of Medical Research, Jan 2026. URL: https://doi.org/10.1186/s40001-025-03655-4, doi:10.1186/s40001-025-03655-4. This article has 0 citations and is from a peer-reviewed journal.
id: P00720
gene_symbol: E
product_type: PROTEIN
status: DRAFT
taxon:
id: NCBITaxon:10665
label: Enterobacteria phage T4
description: >-
T4 endolysin (gene E product, also known as T4 lysozyme) is a GH24 family muramidase
that catalyzes hydrolysis of the beta-1,4-glycosidic bond between N-acetylmuramic acid
and N-acetylglucosamine in bacterial peptidoglycan. The enzyme functions in the periplasm
after holin-mediated membrane permeabilization, degrading the peptidoglycan layer to enable
viral release during the lytic cycle. It is one of the most extensively studied enzymes,
with over 700 crystal structures deposited in the PDB. The catalytic mechanism involves a
conserved Glu11-Asp20-Thr26 triad operating via an inverting glycosidase mechanism.
existing_annotations:
- term:
id: GO:0003796
label: lysozyme activity
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
T4 endolysin has well-established lysozyme (muramidase) activity, catalyzing hydrolysis of
the beta-1,4-glycosidic bond between N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG)
in peptidoglycan [PMID:4865643]. The enzyme belongs to glycosyl hydrolase family 24 (GH24)
with a conserved catalytic triad (Glu11-Asp20-Thr26). This IEA annotation is consistent with
extensive experimental evidence.
action: ACCEPT
reason: >-
Core molecular function of the enzyme. Lysozyme activity is definitively established by
biochemical characterization [PMID:4865643] and structural studies showing substrate binding
and catalytic mechanism [PMID:8266098]. The deep research review confirms T4 endolysin acts as
an endo-acetylmuramidase cleaving glycosidic bonds at muramic acid residues.
additional_reference_ids:
- PMID:4865643
- PMID:8266098
supported_by:
- reference_id: PMID:1201752
supporting_text: "T4 acts as an endo-acetylmuramidase capable of cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains."
- term:
id: GO:0003824
label: catalytic activity
evidence_type: IEA
original_reference_id: GO_REF:0000043
review:
summary: >-
General parent term for catalytic activity. T4 lysozyme is unequivocally a catalyst,
but this term is very general and adds little information beyond the more specific
lysozyme activity annotation.
action: ACCEPT
reason: >-
While redundant with more specific terms, this IEA annotation from keyword mapping is
not incorrect. It serves as a catch-all parent term. The more informative GO:0003796
(lysozyme activity) provides the specific function.
- term:
id: GO:0009253
label: peptidoglycan catabolic process
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
T4 lysozyme degrades peptidoglycan by hydrolyzing glycosidic bonds in the polysaccharide
backbone. This catabolic activity is central to its biological role in enabling phage release
by degrading the host cell wall.
action: ACCEPT
reason: >-
Core biological process annotation. The enzyme directly catalyzes peptidoglycan degradation,
which is its primary biological function. UniProt states it "degrades host peptidoglycans"
[ECO:0000255|HAMAP-Rule:MF_04110]. The deep research confirms the enzyme "directly hydrolyzes
peptidoglycan, weakening the sacculus."
supported_by:
- reference_id: PMID:1201752
supporting_text: "Lysozyme from bacteriophage T4 was found to digest a soluble, uncrosslinked peptidoglycan"
- term:
id: GO:0016787
label: hydrolase activity
evidence_type: IEA
original_reference_id: GO_REF:0000043
review:
summary: >-
General parent term for hydrolase activity. T4 lysozyme catalyzes hydrolysis of glycosidic bonds,
so this parent term is correct but non-specific.
action: ACCEPT
reason: >-
Correct parent term mapping. The enzyme is indeed a hydrolase (EC 3.2.1.17). While more
specific terms are available (lysozyme activity, hydrolase activity acting on glycosyl bonds),
this general IEA annotation is not wrong.
- term:
id: GO:0016798
label: hydrolase activity, acting on glycosyl bonds
evidence_type: IEA
original_reference_id: GO_REF:0000043
review:
summary: >-
T4 lysozyme hydrolyzes beta-1,4 glycosidic bonds between NAM and NAG residues. This term
correctly captures the enzymatic mechanism at an intermediate level of specificity.
action: ACCEPT
reason: >-
Appropriate parent term. T4 lysozyme cleaves glycosidic bonds as established by its EC
number (3.2.1.17) and detailed structural/biochemical studies. This is more informative
than generic hydrolase activity while being less specific than lysozyme activity.
- term:
id: GO:0016998
label: cell wall macromolecule catabolic process
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
Peptidoglycan is a major cell wall macromolecule in bacteria, and T4 lysozyme degrades it.
This term is a parent of peptidoglycan catabolic process.
action: ACCEPT
reason: >-
Correct parent term from InterPro mapping. The enzyme catabolizes peptidoglycan, which is
the major structural component of bacterial cell walls. This is slightly less specific than
GO:0009253 (peptidoglycan catabolic process) but correctly captures the biological process.
- term:
id: GO:0030430
label: host cell cytoplasm
evidence_type: IEA
original_reference_id: GO_REF:0000044
review:
summary: >-
T4 lysozyme accumulates in the host cell cytoplasm during viral replication before being
released to the periplasm via holin-mediated membrane permeabilization.
action: ACCEPT
reason: >-
Correct subcellular localization. UniProt states: "The endolysin is cytoplasmic, but can reach
the periplasmic space with the help of the holins which disrupt the host cell membrane." The
protein is synthesized and accumulates in the cytoplasm, making this a valid localization annotation.
supported_by:
- reference_id: file:BPT4/E/E-deep-research-falcon.md
supporting_text: "T4 E endolysin is produced late, accumulates in the cytoplasm, and acts in the periplasm once holin T forms pores"
- term:
id: GO:0031640
label: killing of cells of another organism
evidence_type: IEA
original_reference_id: GO_REF:0000043
review:
summary: >-
This term describes "killing of cells of another organism" and was mapped from UniProt keywords.
While T4 lysozyme activity does lead to host cell death, this framing is questionable for a
phage-encoded enzyme that kills its own host cell as part of the viral life cycle.
action: MARK_AS_OVER_ANNOTATED
reason: >-
This term is typically used for organisms with distinct identities where one kills cells of
another (e.g., immune cells killing pathogens, antimicrobial peptides). For a phage enzyme,
the host bacterium IS the organism in which the phage replicates - the relationship is more
accurately described as "viral release from host cell" rather than killing another organism's
cells. The term GO:0044659 (viral release from host cell by cytolysis) more accurately captures
this biological context. This appears to be an over-annotation arising from keyword mapping.
- term:
id: GO:0042742
label: defense response to bacterium
evidence_type: IEA
original_reference_id: GO_REF:0000043
review:
summary: >-
This term describes "reactions triggered in response to the presence of a bacterium that act
to protect the cell or organism." This is fundamentally inappropriate for T4 lysozyme - a
bacteriophage is not defending against bacteria; it is parasitizing them.
action: REMOVE
reason: >-
This is a clear case of inappropriate SPKW (SwissProt Keyword) over-annotation. The term
GO:0042742 is designed for eukaryotic innate immunity and antimicrobial defense systems
(e.g., lysozyme in tears defending against bacterial infection). T4 lysozyme is NOT a
defense response - it is an offensive viral mechanism to lyse the host cell for progeny
release. The phage does not "defend" against bacteria; it attacks and destroys them as
part of its reproductive cycle. The correct process annotation is GO:0044659 (viral release
from host cell by cytolysis). This annotation likely arose from the "Lysozyme" keyword being
inappropriately linked to defense responses, without considering that phage lysozymes have
the opposite biological context from eukaryotic lysozymes.
- term:
id: GO:0044659
label: viral release from host cell by cytolysis
evidence_type: IEA
original_reference_id: GO_REF:0000104
review:
summary: >-
T4 lysozyme is an essential component of the phage lysis system, working with holins and
spanins to achieve programmed host cell lysis and release of mature virions.
action: ACCEPT
reason: >-
Core biological process annotation. This accurately describes the primary biological role of
T4 lysozyme. The deep research review states: "T4 E endolysin is produced late, accumulates
in the cytoplasm, and acts in the periplasm once holin T forms pores; it directly hydrolyzes
peptidoglycan, weakening the sacculus. Subsequent spanin action disrupts the outer membrane,
completing lysis in Gram-negative hosts." UniProt confirms it "participates with the holin and
spanin proteins in the sequential events which lead to the programmed host cell lysis releasing
the mature viral particles."
supported_by:
- reference_id: file:BPT4/E/E-deep-research-falcon.md
supporting_text: "Endolysin E functions in the periplasm following holin T pore formation; spanins mediate outer-membrane fusion/rupture to complete host-cell lysis in Gram-negative bacteria"
- term:
id: GO:0003796
label: lysozyme activity
evidence_type: IDA
original_reference_id: PMID:4865643
review:
summary: >-
Tsugita and Inouye (1968) purified T4 lysozyme and characterized its catalytic activity.
This is the foundational experimental paper establishing the biochemical function.
action: ACCEPT
reason: >-
Core experimental evidence for the primary molecular function. PMID:4865643 is cited in
UniProt for the EC number assignment (EC=3.2.1.17) and catalytic activity characterization.
This IDA annotation is well-supported by direct biochemical assays.
supported_by:
- reference_id: PMID:4865643
supporting_text: "Purification of bacteriophage T4 lysozyme"
full_text_unavailable: true
- term:
id: GO:0003796
label: lysozyme activity
evidence_type: IDA
original_reference_id: PMID:1201752
review:
summary: >-
Mirelman et al. (1975) characterized the substrate specificity of T4 lysozyme, demonstrating
it acts as an endo-acetylmuramidase cleaving glycosidic bonds at peptide-substituted muramic
acid residues.
action: ACCEPT
reason: >-
Important experimental evidence providing mechanistic detail on substrate specificity. The
paper directly demonstrates the muramidase activity and substrate requirements.
supported_by:
- reference_id: PMID:1201752
supporting_text: "T4 acts as an endo-acetylmuramidase capable of cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains."
- term:
id: GO:0003796
label: lysozyme activity
evidence_type: IDA
original_reference_id: PMID:4582731
review:
summary: >-
PMID:4582731 (Inouye et al. 1973) is titled "Bacteriophage T7 lysozyme is an
N-acetylmuramyl-L-alanine amidase" and describes T7 lysozyme, NOT T4 lysozyme. This
reference is incorrectly attributed to the T4 gene E product. The annotation itself
(GO:0003796 lysozyme activity) is correct for T4 lysozyme based on other evidence,
but this specific annotation line should be removed due to incorrect reference.
action: ACCEPT
reason: >-
While this specific annotation line has an incorrect reference (PMID:4582731 is about T7 lysozyme,
not T4 lysozyme), the GO term itself (GO:0003796 lysozyme activity) is unequivocally correct
for T4 lysozyme based on extensive other evidence. The term should be ACCEPT because lysozyme
activity is the core function of this enzyme. However, note that this specific reference
attribution is erroneous - T4 and T7 lysozymes are unrelated enzymes with different catalytic
mechanisms (T4 is a muramidase cleaving glycosidic bonds; T7 is an amidase cleaving peptide bonds).
The annotation itself should remain, but the reference should be corrected in the source database.
supported_by:
- reference_id: PMID:4582731
supporting_text: "Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine amidase"
references:
- id: GO_REF:0000002
title: Gene Ontology annotation through association of InterPro records with GO terms
findings: []
- id: GO_REF:0000043
title: Gene Ontology annotation based on UniProtKB/Swiss-Prot keyword mapping
findings:
- statement: Some keywords like "Lysozyme" lead to inappropriate annotations for phage enzymes (e.g., defense response to bacterium)
- id: GO_REF:0000044
title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping
findings: []
- id: GO_REF:0000104
title: Electronic Gene Ontology annotations created by transferring manual GO annotations between related proteins based on shared sequence features
findings: []
- id: GO_REF:0000120
title: Combined Automated Annotation using Multiple IEA Methods
findings: []
- id: PMID:1201752
title: Studies on the specificity of action of bacteriophage T4 lysozyme.
findings:
- statement: T4 lysozyme acts as an endo-acetylmuramidase
supporting_text: "T4 acts as an endo-acetylmuramidase capable of cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains."
- statement: Cleaves glycosidic bonds only at muramic acid residues substituted with peptide side-chains
supporting_text: "cleaving glycosidic bonds only at muramic acid residues that are substituted with peptide side-chains"
- id: PMID:4582731
title: Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine amidase.
findings:
- statement: This paper characterizes T7 lysozyme, not T4 lysozyme - incorrectly cited for T4 gene E
supporting_text: "Bacteriophage T7 lysozyme is an N-acetylmuramyl-L-alanine amidase"
- statement: T7 lysozyme is an amidase (cleaves peptide-glycan bond), distinct from T4 muramidase
supporting_text: "N-acetylmuramyl-L-alanine amidase"
- id: PMID:4865643
title: Purification of bacteriophage T4 lysozyme.
full_text_unavailable: true
findings: []
- id: PMID:8266098
title: A covalent enzyme-substrate intermediate with saccharide distortion in a mutant T4 lysozyme.
findings:
- statement: Structural evidence for catalytic mechanism
supporting_text: "The mutation of threonine 26 to glutamic acid in the active site cleft of phage T4 lysozyme (T4L) produced an enzyme that cleaved the cell wall of Escherichia coli but left the product covalently bound to the enzyme"
- statement: T26H mutant traps covalent intermediate
supporting_text: "analysis of its structure showed a covalent linkage between the product and the newly introduced glutamic acid 26. The covalently linked sugar ring was substantially distorted"
- id: PMID:22389108
title: Protein determinants of phage T4 lysis inhibition.
findings:
- statement: Function of endolysin in lysis pathway
supporting_text: "lysis inhibition in bacteriophage T4 infections occurs when the RI antiholin inhibits the lethal hole-forming function of the T holin"
- statement: Interaction with holin and antiholin system
supporting_text: "the interaction of the soluble domains of these two proteins within the periplasm was necessary for lysis inhibition...Incubation of RI with T inhibits this aggregation and results in a complex of equimolar T and RI content"
core_functions:
- molecular_function:
id: GO:0003796
label: lysozyme activity
description: >-
Definitive core function established by biochemical characterization [PMID:4865643],
substrate specificity studies [PMID:1201752], and extensive structural analysis.
T4 lysozyme is a GH24 muramidase that cleaves the beta-1,4 glycosidic bond between
NAM and NAG in peptidoglycan using a Glu11-Asp20-Thr26 catalytic triad.
directly_involved_in:
- id: GO:0044659
label: viral release from host cell by cytolysis
locations:
- id: GO:0030430
label: host cell cytoplasm
proposed_new_terms: []
suggested_questions:
- question: Is there any evidence for substrate specificity differences between T4 lysozyme and other GH24 lysozymes?
- question: Are there documented cases of T4 lysozyme activity outside the lysis pathway context?
suggested_experiments: []