UbiE (PA5063) is a bifunctional SAM-dependent C-methyltransferase that catalyzes the penultimate C-methylation step in both ubiquinone and menaquinone biosynthesis in P. aeruginosa. In the ubiquinone pathway, it converts 2-polyprenyl-6-methoxy-1,4-benzoquinol to 2-polyprenyl-3-methyl-6-methoxy-1,4-benzoquinol (EC 2.1.1.201); in the menaquinone pathway, it converts demethylmenaquinol to menaquinol (EC 2.1.1.163). Both ubiquinone-9 and menaquinone are essential electron carriers in the branched respiratory chain of P. aeruginosa, supporting aerobic respiration and anaerobic denitrification (PMID:32409583). The enzyme belongs to the class I SAM-binding methyltransferase superfamily (UbiE/COQ5 family). No direct experimental characterization exists in P. aeruginosa; function is inferred from orthologs in E. coli (PMID:9045837), S. cerevisiae COQ5 (PMID:9083049), and mycobacterial MenG (PMID:36417754).
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
|
GO:0006744
ubiquinone biosynthetic process
|
IBA
GO_REF:0000033 |
ACCEPT |
Summary: IBA annotation for ubiquinone biosynthetic process, inferred from phylogenetic trees. UbiE catalyzes the C-methylation of 2-polyprenyl-6-methoxy-1,4-benzoquinol in the ubiquinone pathway. This is a core function of the UbiE/COQ5 family, well established in E. coli (PMID:9045837) and yeast (PMID:9083049).
Reason: Core biosynthetic process. UbiE is directly involved in ubiquinone biosynthesis as the C-methyltransferase for the penultimate step. The BioReason SFT trace (ubiE-deep-research-bioreason-sft.md) also identifies menaquinone headgroup maturation as a core function, consistent with this annotation.
Supporting Evidence:
PMID:9045837
Strains of Escherichia coli with mutations in the ubiE gene are not able to catalyze the carbon methylation reaction in the biosynthesis of ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid quinone components of the respiratory electron transport chain
file:PSEAE/ubiE/ubiE-deep-research-bioreason-sft.md
A SAM-dependent C-methyltransferase in Pseudomonas aeruginosa that installs the C-3 methyl group on demethylated menaquinone (and related demethylated quinone intermediates), thereby advancing menaquinone headgroup maturation
|
|
GO:0008425
2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
|
IBA
GO_REF:0000033 |
ACCEPT |
Summary: IBA annotation for the specific molecular function in the ubiquinone pathway (EC 2.1.1.201). This is one of the two core catalytic activities of UbiE, converting DDMQH2 to DMQH2 via SAM-dependent C-methylation. Well supported by ortholog data from E. coli and yeast COQ5.
Reason: Core molecular function. This is one of the two specific methyltransferase activities of UbiE, directly corresponding to EC 2.1.1.201.
Supporting Evidence:
PMID:9083049
Coq5p is required for the C-methyltransferase step that converts 2-methoxy-6-polyprenyl-1, 4-benzoquinone to 2-methoxy-5-methyl-6-polyprenyl-1,4-benzoquinone
|
|
GO:0006744
ubiquinone biosynthetic process
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: IEA annotation for ubiquinone biosynthetic process from UniRule/UniPathway. Redundant with the IBA annotation above but independently supported. Correct annotation for a core biosynthetic function.
Reason: Correct and redundant with phylogenetic inference. UniRule UR000030800 (HAMAP MF_01813) appropriately assigns this term to the UbiE/COQ5 family.
|
|
GO:0008168
methyltransferase activity
|
IEA
GO_REF:0000002 |
MODIFY |
Summary: InterPro2GO annotation from IPR004033 (UbiE/COQ5 methyltransferase) and IPR023576 (conserved site). The general methyltransferase activity term is correct but overly broad. More specific terms (GO:0008425 and GO:0043770) are already annotated and better represent the actual catalytic activities.
Reason: Correct but too general. The more specific terms GO:0008425 (2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity) and GO:0043770 (demethylmenaquinone methyltransferase activity) are already present and more informative.
|
|
GO:0008425
2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: IEA annotation for the ubiquinone-pathway methyltransferase activity from UniRule. Correct and specific, corresponding to EC 2.1.1.201. Redundant with the IBA annotation.
Reason: Correct specific molecular function. Independently supported by both UniRule/Rhea and phylogenetic inference.
|
|
GO:0009060
aerobic respiration
|
IEA
GO_REF:0000104 |
KEEP AS NON CORE |
Summary: IEA annotation for aerobic respiration. Ubiquinone is indispensable for the aerobic electron transport chain in P. aeruginosa (PMID:6774977), so ubiE is required for aerobic respiration. However, this is an indirect downstream consequence of ubiquinone biosynthesis rather than a direct function of ubiE. The enzyme catalyzes a biosynthetic step; it does not directly participate in electron transport.
Reason: While correct that UbiE activity is required for aerobic respiration (because ubiquinone is essential for the respiratory chain), this annotation describes a downstream physiological consequence rather than the direct molecular role. The core annotation is the biosynthetic process.
Supporting Evidence:
PMID:6774977
Complete removal of ubiquinone performed by extracting the lyophilized membrane particles with n-pentane containing acetone resulted in complete loss of all oxidase activities for glucose, gluconate, malate, succinate, and NADH
|
|
GO:0009234
menaquinone biosynthetic process
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: IEA annotation for menaquinone biosynthetic process from UniRule/UniPathway. UbiE catalyzes the final step in menaquinone headgroup maturation, converting demethylmenaquinol to menaquinol. This is the second of the two core biosynthetic pathways involving UbiE.
Reason: Core biosynthetic process. UbiE/MenG directly catalyzes the terminal C-methylation step in menaquinone biosynthesis, well established in E. coli and mycobacteria.
Supporting Evidence:
PMID:9045837
Strains of Escherichia coli with mutations in the ubiE gene are not able to catalyze the carbon methylation reaction in the biosynthesis of ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid quinone components of the respiratory electron transport chain
|
|
GO:0042181
ketone biosynthetic process
|
IEA
GO_REF:0000117 |
MARK AS OVER ANNOTATED |
Summary: ARBA-assigned annotation for ketone biosynthetic process. Ubiquinone and menaquinone are quinones (not ketones per se, although quinones contain carbonyl groups). This is an overly general and somewhat misleading parent term. The specific pathway terms (GO:0006744 ubiquinone biosynthetic process and GO:0009234 menaquinone biosynthetic process) are already present and far more informative.
Reason: Technically not wrong (quinones contain ketone-like carbonyl groups), but this is an uninformative high-level grouping term that obscures the actual biology. The specific quinone biosynthesis terms already capture the biology precisely.
|
|
GO:0043770
demethylmenaquinone methyltransferase activity
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: IEA annotation for the menaquinone-pathway methyltransferase activity (EC 2.1.1.163) from UniRule. This is the second of the two core catalytic activities of UbiE, converting demethylmenaquinol to menaquinol. Well supported by ortholog data, including direct enzymatic characterization of mycobacterial MenG (PMID:36417754).
Reason: Core molecular function. This is the specific methyltransferase activity for the menaquinone pathway, directly corresponding to EC 2.1.1.163.
Supporting Evidence:
PMID:36417754
Partially purified recombinant protein catalyzed the formation of MK from demethylmenaquinone (DMK), although the activity of the recombinant protein was low and appeared to require a cofactor or intact membrane structure for activity
|
Q: Does P. aeruginosa ubiE have a preference for the ubiquinone versus menaquinone pathway substrate? The relative importance of each pathway may differ from E. coli given that UQ9 is the dominant quinone in P. aeruginosa.
Suggested experts: Fabien Pierrel, Ludovic Pelosi
Q: Is there any direct experimental evidence for ubiE function in P. aeruginosa (e.g., gene knockout, complementation, or enzymatic assay)?
Experiment: Generate a clean deletion of PA5063 (ubiE) in P. aeruginosa PAO1 using allelic exchange. Assess growth phenotype on aerobic and anaerobic (denitrifying) media. Analyze quinone profiles (UQ9, MK, DMK) by HPLC-MS. Complement with wild-type ubiE expressed from a plasmid.
Hypothesis: P. aeruginosa ubiE knockout would be deficient in both ubiquinone and menaquinone biosynthesis, resulting in growth defects under both aerobic and anaerobic conditions.
Type: gene knockout and metabolite profiling
I begin with the InterPro architecture. The core of the protein is defined by IPR004033 (UbiE/COQ5 methyltransferase family) spanning residues 21–255, with three overlapping entries across 21–255/25–255/30–255, indicating a canonical UbiE/COQ5 fold. This family assignment is embedded within IPR029063 (S-adenosyl-L-methionine-dependent methyltransferase superfamily, residues 18–253 and 36–256), which denotes a Rossmann-like class I SAM-dependent methyltransferase scaffold. Two conserved sites punctuate this core: IPR023576 at residues 41–56 and residues 163–177. These motifs typically organize the SAM-binding pocket and the catalytic environment that stabilizes the transition state for electrophilic methyl transfer to a carbon center. The ordered layout—superfamily Rossmann-like SAM-MTase scaffold with UbiE/COQ5 family determinants and two conserved catalytic/ligand-binding sites—causes a methyltransferase chemistry that uses S-adenosyl-L-methionine (SAM) to methylate a quinone ring at a defined carbon.
From this architecture, the molecular function resolves to a SAM-dependent quinone C-methylation. The UbiE/COQ5 family is specialized for methylating prenylated quinones at the C-3 position of demethylated intermediates. The presence of the UbiE/COQ5 family signatures together with the conserved sites that coordinate SAM and the quinone substrate supports the specific activity formalized as GO:0008924 3-demethylubiquinone methyltransferase activity. The same catalytic logic applies to demethylmenaquinone substrates in some bacteria, because the reaction is a carbon–carbon bond-forming methylation of a quinone headgroup; thus the enzyme can act on 3-demethylmenaquinone as well as 3-demethylubiquinone.
This chemistry feeds directly into quinone biosynthesis. By installing the C-3 methyl group, the enzyme completes the headgroup maturation of demethylmenaquinone, advancing it toward functional menaquinone. This places the protein in the biological process GO:0009234 menaquinone biosynthetic process. In organisms that use both ubiquinone and menaquinone pathways, UbiE/COQ5-like enzymes often exhibit substrate breadth governed by active-site determinants encoded in the family core; here, the UbiE/COQ5 family signatures and conserved sites are consistent with catalysis on demethylmenaquinone as part of the menaquinone branch.
The cellular context follows from the pathway organization. Quinone biosynthesis in Pseudomonas aeruginosa proceeds at the cytoplasmic face of the inner (plasma) membrane, where hydrophobic prenylated intermediates partition. UbiE/COQ5-like enzymes typically assemble into multi-enzyme metabolons with other quinone biosynthetic proteins to channel hydrophobic intermediates. This supports a residence within a GO:0032991 protein-containing complex. Given the membrane-proximal nature of the substrates and the need for efficient handoff among pathway enzymes, the most plausible physical setting is at the bacterial plasma membrane, where the enzyme associates peripherally with the membrane-resident biosynthetic machinery.
Mechanistically, the enzyme binds SAM in the Rossmann-like pocket formed by the class I methyltransferase superfamily scaffold (IPR029063) and positions 3-demethylmenaquinone (or 3-demethylubiquinone) in a hydrophobic channel shaped by the UbiE/COQ5 family core (IPR004033). The conserved sites at residues 41–56 and 163–177 stabilize the transition state for electrophilic substitution at C-3 of the quinone headgroup, transferring the methyl from SAM to yield the methylated quinone and S-adenosyl-L-homocysteine. Within a membrane-associated quinone biosynthetic complex, it likely receives the demethylated quinone from upstream methyltransferases and passes the product to downstream tailoring or modification steps.
This model predicts specific interaction partners. Association with 4-hydroxybenzoate-octaprenyl transferase positions the enzyme downstream of prenylation, while proximity to 3-demethylubiquinone 3-methyltransferase and Ubiquinone biosynthetic protein UbiB would organize a quinone-biosynthesis metabolon at the membrane. Contacts with NAD(P)H quinone oxidoreductase and probable NAD(P)H dehydrogenases could couple redox poise to quinone maturation, while the Trp repressor binding protein WrbA may buffer reactive oxygen species generated during quinone processing. The presence of a hypothetical protein partner suggests accessory scaffolds or adaptors that stabilize the complex. Together, these interactions would create a membrane-proximal assembly that channels hydrophobic intermediates and coordinates methylation with the rest of the quinone pathway.
## Functional Summary
A SAM-dependent C-methyltransferase in Pseudomonas aeruginosa that installs the C-3 methyl group on demethylated menaquinone (and related demethylated quinone intermediates), thereby advancing menaquinone headgroup maturation within a membrane-associated quinone-biosynthesis assembly. The enzyme uses a Rossmann-like class I methyltransferase core to bind S-adenosyl-L-methionine and a UbiE/COQ5-specific pocket to position the quinone ring for electrophilic methyl transfer, likely operating at the cytoplasmic face of the inner membrane as part of a multi-enzyme complex that channels hydrophobic intermediates between pathway steps.
## UniProt Summary
Menaquinone biosynthesis.
## InterPro Domains
- IPR029063: S-adenosyl-L-methionine-dependent methyltransferase superfamily (homologous_superfamily) [18-253]
- IPR004033: UbiE/COQ5 methyltransferase (family) [21-255]
- IPR004033: UbiE/COQ5 methyltransferase (family) [25-255]
- IPR004033: UbiE/COQ5 methyltransferase (family) [30-255]
- IPR029063: S-adenosyl-L-methionine-dependent methyltransferase superfamily (homologous_superfamily) [36-256]
- IPR023576: UbiE/COQ5 methyltransferase, conserved site (conserved_site) [41-56]
- IPR023576: UbiE/COQ5 methyltransferase, conserved site (conserved_site) [163-177]
## GO Term Predictions
### Molecular Function
### Biological Process
### Cellular Component
UbiE is a bifunctional SAM-dependent C-methyltransferase that catalyzes the penultimate C-methylation step in both ubiquinone (coenzyme Q) and menaquinone (vitamin K2) biosynthesis. The E. coli ortholog was originally identified by Lee et al. (1997), who demonstrated that ubiE mutants fail to catalyze the C-methylation reaction in both pathways PMID:9045837. The same study showed that disruption of ubiE causes accumulation of 2-octaprenyl-6-methoxy-1,4-benzoquinone and demethylmenaquinone as predominant intermediates PMID:9045837.
The yeast ortholog COQ5 was independently characterized by Barkovich et al. (1997), who confirmed C-methyltransferase activity and mitochondrial localization PMID:9083049.
The enzyme has two distinct catalytic activities:
In the mycobacterial ortholog (MenG/Rv0558), Pujari et al. (2022) demonstrated that the actual substrate is the reduced form (demethylmenaquinol), as addition of reductants stimulated activity PMID:36417754.
P. aeruginosa uses ubiquinone-9 (coenzyme Q9) as the primary electron carrier in its aerobic respiratory chain. Matsushita et al. (1980) established that ubiquinone is indispensable for respiratory chain function in P. aeruginosa PMID:6774977. The respiratory chain of aerobically grown P. aeruginosa contains coenzyme Q9 along with b-type and c-type cytochromes PMID:6766443.
Critically, Vo et al. (2020) demonstrated that UQ9 is required for anaerobic denitrification in P. aeruginosa, not just aerobic respiration PMID:32409583. This means that both ubiquinone and menaquinone biosynthesis are physiologically relevant in this organism, and the ubiE C-methylation step is essential for both pathways.
E. coli studies showed that menaquinone (but not demethylmenaquinone) is required for certain anaerobic electron transfer pathways. Tyson et al. (1997) demonstrated that ubiE mutants, which produce demethylmenaquinone but not menaquinone, retain some but not all respiratory activity PMID:9325429. This work also established that menaquinone is essential for cytochrome-c-dependent TMAO reductase and Nrf activities.
In E. coli, ubiE is the first gene of an operon containing ubiE-yigP-ubiB PMID:10960098. In P. aeruginosa PAO1, ubiE (PA5063) is similarly located in a genomic context with genes involved in quinone metabolism, identified through the complete genome sequence PMID:10984043.
The protein belongs to the class I SAM-binding methyltransferase superfamily (IPR029063) with a Rossmann-like fold. It contains the UbiE/COQ5 family signature (IPR004033) and conserved sites (IPR023576) that coordinate SAM binding. The protein has 256 amino acids with SAM-binding residues at positions 79, 100, and 128-129 (UniProt annotation by HAMAP rule MF_01813).
P. aeruginosa ubiE (PA5063) encodes a bifunctional SAM-dependent C-methyltransferase that is required for the C-methylation step in both ubiquinone and menaquinone biosynthesis. Both products are essential for the branched respiratory chain of P. aeruginosa, which uses ubiquinone for both aerobic and anaerobic (denitrification) respiration. The protein is annotated entirely by homology transfer (no direct experimental evidence in P. aeruginosa), but the function is strongly supported by experimental characterization of orthologs in E. coli, yeast, and mycobacteria.
Source: ubiE-deep-research-bioreason-sft.md
The BioReason functional summary states:
A SAM-dependent C-methyltransferase in Pseudomonas aeruginosa that installs the C-3 methyl group on demethylated menaquinone (and related demethylated quinone intermediates), thereby advancing menaquinone headgroup maturation within a membrane-associated quinone-biosynthesis assembly. The enzyme uses a Rossmann-like class I methyltransferase core to bind S-adenosyl-L-methionine and a UbiE/COQ5-specific pocket to position the quinone ring for electrophilic methyl transfer, likely operating at the cytoplasmic face of the inner membrane as part of a multi-enzyme complex that channels hydrophobic intermediates between pathway steps.
The core catalytic identity -- SAM-dependent C-methyltransferase that methylates demethylated quinone intermediates -- is correct and well-stated. The mechanistic detail about the Rossmann-like class I methyltransferase core and SAM binding is accurate. The description of the UbiE/COQ5-specific pocket positioning the quinone ring for electrophilic methyl transfer matches the known biochemistry.
However, there are notable issues:
Menaquinone bias: The summary heavily emphasizes menaquinone biosynthesis while only parenthetically mentioning "related demethylated quinone intermediates." In reality, UbiE is well established as a bifunctional enzyme acting in BOTH ubiquinone and menaquinone biosynthesis (PMID:9045837). The ubiquinone pathway role (EC 2.1.1.201, GO:0008425) is equally important, especially in P. aeruginosa where UQ9 is the dominant quinone (PMID:6774977, PMID:32409583). The omission of explicit mention of ubiquinone biosynthesis is a significant gap.
Speculative complex formation: The summary claims the enzyme operates "as part of a multi-enzyme complex that channels hydrophobic intermediates." While metabolon formation has been hypothesized for quinone biosynthesis, direct evidence for a UbiE-containing complex in bacteria is limited. The thinking trace goes further to name specific interaction partners (UbiB, quinone oxidoreductases, WrbA), which are speculative.
Cellular localization speculation: The claim of operating at "the cytoplasmic face of the inner membrane" is plausible but unsupported by direct evidence. UbiE is a soluble protein; the membrane association, if any, would be through interaction with hydrophobic substrates or other membrane-associated proteins.
No mention of aerobic/anaerobic respiration context: The physiological significance of UbiE in supporting both aerobic respiration and anaerobic denitrification in P. aeruginosa (PMID:32409583) is not discussed.
Correct GO term cited but rare: The thinking trace cites GO:0008924 (3-demethylubiquinone methyltransferase activity), which is not the standard term used in current annotations. The curated annotations use GO:0008425 (2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity) and GO:0043770 (demethylmenaquinone methyltransferase activity).
Comparison with interpro2go:
The InterPro2GO annotation (GO_REF:0000002) assigns GO:0008168 (methyltransferase activity) based on IPR004033 and IPR023576. This is a very broad, non-specific assignment. BioReason substantially improves on interpro2go by:
However, BioReason does NOT simply recapitulate interpro2go. It adds genuine biological insight about the catalytic mechanism and pathway context. The main weakness compared to the curated review is the incomplete coverage of the ubiquinone pathway role and the speculative claims about complex formation and membrane association. The interpro2go annotation, despite being broad, at least avoids these speculative errors.
The thinking trace systematically walks through InterPro domain architecture, which is methodologically sound. The identification of IPR004033 (UbiE/COQ5 family), IPR029063 (SAM-dependent methyltransferase superfamily), and IPR023576 (conserved sites) is accurate and well-reasoned. The deduction of SAM-dependent quinone C-methylation from these domains is logical and correct.
The trace becomes speculative when it infers protein complex membership (GO:0032991) and specific interaction partners. The claim that the enzyme "likely receives the demethylated quinone from upstream methyltransferases and passes the product to downstream tailoring or modification steps" is a reasonable hypothesis but presented with unwarranted confidence. The naming of specific interaction partners (4-hydroxybenzoate-octaprenyl transferase, UbiB, WrbA, NAD(P)H quinone oxidoreductase) appears to be drawn from STRING or similar interaction databases and is presented as if it were established biology.
The GO term predictions section is empty (no MF, BP, or CC predictions listed), which is unusual. The functional summary and thinking trace contain the BioReason reasoning, while the GO term predictions were apparently not populated for this SFT entry.
id: Q9HUC0
gene_symbol: ubiE
product_type: PROTEIN
status: COMPLETE
taxon:
id: NCBITaxon:208964
label: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM
14847 / LMG 12228 / 1C / PRS 101 / PAO1)
description: >-
UbiE (PA5063) is a bifunctional SAM-dependent C-methyltransferase that catalyzes
the penultimate C-methylation step in both ubiquinone and menaquinone biosynthesis
in P. aeruginosa. In the ubiquinone pathway, it converts 2-polyprenyl-6-methoxy-1,4-benzoquinol
to 2-polyprenyl-3-methyl-6-methoxy-1,4-benzoquinol (EC 2.1.1.201); in the
menaquinone pathway, it converts demethylmenaquinol to menaquinol (EC 2.1.1.163).
Both ubiquinone-9 and menaquinone are essential electron carriers in the branched
respiratory chain of P. aeruginosa, supporting aerobic respiration and anaerobic
denitrification (PMID:32409583). The enzyme belongs to the class I SAM-binding
methyltransferase superfamily (UbiE/COQ5 family). No direct experimental characterization
exists in P. aeruginosa; function is inferred from orthologs in E. coli (PMID:9045837),
S. cerevisiae COQ5 (PMID:9083049), and mycobacterial MenG (PMID:36417754).
references:
- id: PMID:9045837
title: A C-methyltransferase involved in both ubiquinone and menaquinone biosynthesis;
isolation and identification of the Escherichia coli ubiE gene
findings:
- statement: >-
E. coli ubiE encodes a C-methyltransferase required for C-methylation in
both ubiquinone and menaquinone biosynthesis.
supporting_text: >-
Strains of Escherichia coli with mutations in the ubiE gene are not able
to catalyze the carbon methylation reaction in the biosynthesis of
ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid
quinone components of the respiratory electron transport chain
- statement: >-
ubiE mutants accumulate demethylmenaquinone and 2-octaprenyl-6-methoxy-1,4-benzoquinone.
supporting_text: >-
E. coli strains containing either the disruption or the point mutation in
ubiE accumulated 2-octaprenyl-6-methoxy-1,4-benzoquinone and demethylmenaquinone
as predominant intermediates
- statement: >-
UbiE contains SAM-binding motifs common to SAM-dependent methyltransferases.
supporting_text: >-
we evaluated the amino acid sequences encoded by open reading frames
located in this region for the presence of sequence motifs common to a
wide variety of S-adenosyl-L-methionine-dependent methyltransferases
- id: PMID:9083049
title: Characterization of the COQ5 gene from Saccharomyces cerevisiae. Evidence
for a C-methyltransferase in ubiquinone biosynthesis
findings:
- statement: >-
Yeast COQ5 has 44% identity to E. coli UbiE and is required for a
C-methyltransferase step in ubiquinone biosynthesis.
supporting_text: >-
with 44% sequence identity over 262 amino acids to UbiE, which is
required for a C-methyltransferase step in the Q and menaquinone biosynthetic
pathways in Escherichia coli. Both the ubiE and COQ5 coding sequences contain
sequence motifs common to a wide variety of S-adenosyl-L-methionine-dependent
methyltransferases
- id: PMID:36417754
title: Mycobacterial MenG; Partial Purification, Characterization, and Inhibition
findings:
- statement: >-
Mycobacterial MenG (Rv0558) is a SAM/demethylmenaquinol methyltransferase
that complements E. coli ubiE deletion.
supporting_text: >-
Heterologous expression of Rv0558 complemented an ubiE (the quinone
C-methyltransferase involved in ubiquinone and menaquinone synthesis) deletion
in Escherichia coli, and expression in a wild-type E. coli strain increased
quinone C-methyltransferase specific activity by threefold
- statement: >-
The reduced form demethylmenaquinol, not the oxidized demethylmenaquinone,
is the true substrate.
supporting_text: >-
addition of dithiothreitol, dithionite, NADH, or other substrates of
primary dehydrogenases to reaction mixtures containing membrane preparations
stimulated the activity. Thus, these observations strongly suggest that
demethylmenaquinol is the actual substrate of MenG
- id: PMID:10960098
title: Identification of Escherichia coli ubiB, a gene required for the first
monooxygenase step in ubiquinone biosynthesis
findings:
- statement: >-
ubiE is the first gene in an operon with yigP and ubiB in E. coli.
supporting_text: >-
the ubiE gene encodes a C-methyltransferase required for the synthesis of
both CoQ and menaquinone, and it is the 5' gene in an operon containing
ubiE, yigP, and ubiB
- id: PMID:9325429
title: Characterisation of Escherichia coli K-12 mutants defective in formate-dependent
nitrite reduction; essential roles for hemN and the menFDBCE operon
findings:
- statement: >-
In E. coli, menaquinone (but not demethylmenaquinone) is essential for
certain anaerobic electron transfer pathways.
supporting_text: >-
combined data establish that menaquinones are essential for
cytochrome-c-dependent trimethylamine-N-oxide reductase (Tor) and Nrf
activity, but that either menaquinone or ubiquinone, but not
demethylmenaquinone, can transfer electrons to a third
cytochrome-c-dependent electron transfer chain, the periplasmic nitrate
reductase
- id: PMID:32409583
title: The O(2)-independent pathway of ubiquinone biosynthesis is essential for
denitrification in Pseudomonas aeruginosa.
findings:
- statement: >-
Ubiquinone-9 is the major quinone in P. aeruginosa and is required for
anaerobic denitrification.
supporting_text: >-
we established that UQ9 is the major quinone of Pseudomonas aeruginosa
and is required for growth under anaerobic respiration (i.e. denitrification)
- id: PMID:6774977
title: Function of ubiquinone in the electron transport system of Pseudomonas aeruginosa
grown aerobically
findings:
- statement: >-
Ubiquinone is indispensable for the aerobic electron transport chain of
P. aeruginosa.
supporting_text: >-
Complete removal of ubiquinone performed by extracting the lyophilized
membrane particles with n-pentane containing acetone resulted in complete
loss of all oxidase activities for glucose, gluconate, malate, succinate,
and NADH
- id: PMID:6766443
title: Membrane-bound respiratory chain of Pseudomonas aeruginosa grown aerobically
findings:
- statement: >-
Aerobically grown P. aeruginosa uses coenzyme Q9 in its respiratory chain.
supporting_text: >-
The electron transport chain of the gram-negative bacterium Pseudomonas
aeruginosa, grown aerobically, contained a number of primary dehydrogenases
and respiratory components (soluble flavin, bound flavin, coenzyme Q9,
heme b, heme c, and cytochrome o)
- id: PMID:10984043
title: Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic
pathogen
findings: []
- id: GO_REF:0000002
title: Gene Ontology annotation through association of InterPro records with GO terms
findings: []
- id: GO_REF:0000033
title: Annotation inferences using phylogenetic trees
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:0000117
title: Electronic Gene Ontology annotations created by ARBA machine learning models
findings: []
- id: GO_REF:0000120
title: Combined Automated Annotation using Multiple IEA Methods
findings: []
existing_annotations:
# ============================================================
# IBA annotations (phylogenetic inference from PANTHER)
# ============================================================
- term:
id: GO:0006744
label: ubiquinone biosynthetic process
evidence_type: IBA
original_reference_id: GO_REF:0000033
review:
summary: >-
IBA annotation for ubiquinone biosynthetic process, inferred from phylogenetic
trees. UbiE catalyzes the C-methylation of 2-polyprenyl-6-methoxy-1,4-benzoquinol
in the ubiquinone pathway. This is a core function of the UbiE/COQ5 family,
well established in E. coli (PMID:9045837) and yeast (PMID:9083049).
action: ACCEPT
reason: >-
Core biosynthetic process. UbiE is directly involved in ubiquinone biosynthesis
as the C-methyltransferase for the penultimate step. The BioReason SFT trace
(ubiE-deep-research-bioreason-sft.md) also identifies menaquinone headgroup
maturation as a core function, consistent with this annotation.
supported_by:
- reference_id: PMID:9045837
supporting_text: >-
Strains of Escherichia coli with mutations in the ubiE gene are not able
to catalyze the carbon methylation reaction in the biosynthesis of
ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid
quinone components of the respiratory electron transport chain
- reference_id: file:PSEAE/ubiE/ubiE-deep-research-bioreason-sft.md
supporting_text: >-
A SAM-dependent C-methyltransferase in Pseudomonas aeruginosa that
installs the C-3 methyl group on demethylated menaquinone (and related
demethylated quinone intermediates), thereby advancing menaquinone
headgroup maturation
- term:
id: GO:0008425
label: 2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
evidence_type: IBA
original_reference_id: GO_REF:0000033
review:
summary: >-
IBA annotation for the specific molecular function in the ubiquinone pathway
(EC 2.1.1.201). This is one of the two core catalytic activities of UbiE,
converting DDMQH2 to DMQH2 via SAM-dependent C-methylation. Well supported
by ortholog data from E. coli and yeast COQ5.
action: ACCEPT
reason: >-
Core molecular function. This is one of the two specific methyltransferase
activities of UbiE, directly corresponding to EC 2.1.1.201.
supported_by:
- reference_id: PMID:9083049
supporting_text: >-
Coq5p is required for the C-methyltransferase step that converts
2-methoxy-6-polyprenyl-1, 4-benzoquinone to
2-methoxy-5-methyl-6-polyprenyl-1,4-benzoquinone
# ============================================================
# IEA annotations (electronic)
# ============================================================
- term:
id: GO:0006744
label: ubiquinone biosynthetic process
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
IEA annotation for ubiquinone biosynthetic process from UniRule/UniPathway.
Redundant with the IBA annotation above but independently supported.
Correct annotation for a core biosynthetic function.
action: ACCEPT
reason: >-
Correct and redundant with phylogenetic inference. UniRule UR000030800
(HAMAP MF_01813) appropriately assigns this term to the UbiE/COQ5 family.
- term:
id: GO:0008168
label: methyltransferase activity
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
InterPro2GO annotation from IPR004033 (UbiE/COQ5 methyltransferase) and
IPR023576 (conserved site). The general methyltransferase activity term is
correct but overly broad. More specific terms (GO:0008425 and GO:0043770)
are already annotated and better represent the actual catalytic activities.
action: MODIFY
reason: >-
Correct but too general. The more specific terms GO:0008425
(2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity) and
GO:0043770 (demethylmenaquinone methyltransferase activity) are already
present and more informative.
proposed_replacement_terms:
- id: GO:0008425
label: 2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
- id: GO:0043770
label: demethylmenaquinone methyltransferase activity
- term:
id: GO:0008425
label: 2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
IEA annotation for the ubiquinone-pathway methyltransferase activity from
UniRule. Correct and specific, corresponding to EC 2.1.1.201. Redundant
with the IBA annotation.
action: ACCEPT
reason: >-
Correct specific molecular function. Independently supported by both
UniRule/Rhea and phylogenetic inference.
- term:
id: GO:0009060
label: aerobic respiration
evidence_type: IEA
original_reference_id: GO_REF:0000104
review:
summary: >-
IEA annotation for aerobic respiration. Ubiquinone is indispensable for
the aerobic electron transport chain in P. aeruginosa (PMID:6774977), so
ubiE is required for aerobic respiration. However, this is an indirect
downstream consequence of ubiquinone biosynthesis rather than a direct
function of ubiE. The enzyme catalyzes a biosynthetic step; it does not
directly participate in electron transport.
action: KEEP_AS_NON_CORE
reason: >-
While correct that UbiE activity is required for aerobic respiration
(because ubiquinone is essential for the respiratory chain), this
annotation describes a downstream physiological consequence rather than
the direct molecular role. The core annotation is the biosynthetic process.
supported_by:
- reference_id: PMID:6774977
supporting_text: >-
Complete removal of ubiquinone performed by extracting the lyophilized
membrane particles with n-pentane containing acetone resulted in complete
loss of all oxidase activities for glucose, gluconate, malate, succinate,
and NADH
- term:
id: GO:0009234
label: menaquinone biosynthetic process
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
IEA annotation for menaquinone biosynthetic process from UniRule/UniPathway.
UbiE catalyzes the final step in menaquinone headgroup maturation, converting
demethylmenaquinol to menaquinol. This is the second of the two core
biosynthetic pathways involving UbiE.
action: ACCEPT
reason: >-
Core biosynthetic process. UbiE/MenG directly catalyzes the terminal
C-methylation step in menaquinone biosynthesis, well established in
E. coli and mycobacteria.
supported_by:
- reference_id: PMID:9045837
supporting_text: >-
Strains of Escherichia coli with mutations in the ubiE gene are not able
to catalyze the carbon methylation reaction in the biosynthesis of
ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid
quinone components of the respiratory electron transport chain
- term:
id: GO:0042181
label: ketone biosynthetic process
evidence_type: IEA
original_reference_id: GO_REF:0000117
review:
summary: >-
ARBA-assigned annotation for ketone biosynthetic process. Ubiquinone and
menaquinone are quinones (not ketones per se, although quinones contain
carbonyl groups). This is an overly general and somewhat misleading parent
term. The specific pathway terms (GO:0006744 ubiquinone biosynthetic
process and GO:0009234 menaquinone biosynthetic process) are already present
and far more informative.
action: MARK_AS_OVER_ANNOTATED
reason: >-
Technically not wrong (quinones contain ketone-like carbonyl groups),
but this is an uninformative high-level grouping term that obscures the
actual biology. The specific quinone biosynthesis terms already capture
the biology precisely.
- term:
id: GO:0043770
label: demethylmenaquinone methyltransferase activity
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
IEA annotation for the menaquinone-pathway methyltransferase activity
(EC 2.1.1.163) from UniRule. This is the second of the two core catalytic
activities of UbiE, converting demethylmenaquinol to menaquinol. Well
supported by ortholog data, including direct enzymatic characterization of
mycobacterial MenG (PMID:36417754).
action: ACCEPT
reason: >-
Core molecular function. This is the specific methyltransferase activity
for the menaquinone pathway, directly corresponding to EC 2.1.1.163.
supported_by:
- reference_id: PMID:36417754
supporting_text: >-
Partially purified recombinant protein catalyzed the formation of MK
from demethylmenaquinone (DMK), although the activity of the recombinant
protein was low and appeared to require a cofactor or intact membrane
structure for activity
core_functions:
- description: >-
C-methylation of 2-polyprenyl-6-methoxy-1,4-benzoquinol in the ubiquinone
biosynthetic pathway, using SAM as methyl donor
molecular_function:
id: GO:0008425
label: 2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity
directly_involved_in:
- id: GO:0006744
label: ubiquinone biosynthetic process
supported_by:
- reference_id: PMID:9045837
supporting_text: >-
Strains of Escherichia coli with mutations in the ubiE gene are not able
to catalyze the carbon methylation reaction in the biosynthesis of
ubiquinone (coenzyme Q) and menaquinone (vitamin K2), essential isoprenoid
quinone components of the respiratory electron transport chain
- description: >-
C-methylation of demethylmenaquinol in the menaquinone biosynthetic pathway,
using SAM as methyl donor. This is the terminal step in menaquinone headgroup
maturation.
molecular_function:
id: GO:0043770
label: demethylmenaquinone methyltransferase activity
directly_involved_in:
- id: GO:0009234
label: menaquinone biosynthetic process
supported_by:
- reference_id: PMID:36417754
supporting_text: >-
Partially purified recombinant protein catalyzed the formation of MK from
demethylmenaquinone (DMK), although the activity of the recombinant protein
was low and appeared to require a cofactor or intact membrane structure for
activity
suggested_questions:
- question: >-
Does P. aeruginosa ubiE have a preference for the ubiquinone versus menaquinone
pathway substrate? The relative importance of each pathway may differ from E. coli
given that UQ9 is the dominant quinone in P. aeruginosa.
experts:
- Fabien Pierrel
- Ludovic Pelosi
- question: >-
Is there any direct experimental evidence for ubiE function in P. aeruginosa
(e.g., gene knockout, complementation, or enzymatic assay)?
experts: []
suggested_experiments:
- hypothesis: >-
P. aeruginosa ubiE knockout would be deficient in both ubiquinone and menaquinone
biosynthesis, resulting in growth defects under both aerobic and anaerobic conditions.
description: >-
Generate a clean deletion of PA5063 (ubiE) in P. aeruginosa PAO1 using allelic
exchange. Assess growth phenotype on aerobic and anaerobic (denitrifying) media.
Analyze quinone profiles (UQ9, MK, DMK) by HPLC-MS. Complement with wild-type
ubiE expressed from a plasmid.
experiment_type: gene knockout and metabolite profiling