# Heptaprenylglyceryl ether-lipid comparative audit

## Scope and sampled proteins

| Protein | Species/strain | Role and evidential boundary |
|---|---|---|
| P94527 EgsA | Bacillus subtilis 168 | Purified G1PDH capability; native supply to PcrB unresolved |
| O34790 PcrB | Bacillus subtilis 168 | C35 ether-bond enzyme; native carrier-pathway evidence |
| O34993 YvoF | Bacillus subtilis 168 | Acetyl-CoA-dependent O-acetyl loading; membrane-associated activity |
| P54392 YpjA/LhaT | Bacillus subtilis 168 | Lipid-donor-dependent lipoprotein N-acetyl transfer; no identified flippase |
| Q5L3C1 PcrB | Geobacillus kaustophilus HTA426 | C35 step and G1P-bound structure; native downstream capacity unresolved |
| A7X435 PcrB | Staphylococcus aureus Mu3 | Comparative PcrB structure and enzyme capacity; native downstream capacity unresolved |
| A5FJK8 Fjoh_1584 | Flavobacterium johnsoniae UW101 | C20 GGGPS excluded comparator; no C35 carrier-pathway claim |

The module has five roles; four have grounded family descriptors. The required
phosphatase has no established native protein and therefore no defensible family
identifier. PANTHER PTHR43622:SF1 is heterogeneous; its member EgsA requires an
additional G1PDH activity qualification. PcrB is PTHR40029:SF2, YvoF is
PTHR43300:SF6, and LhaT is PTHR40042:SF1 with explicit activity qualification.
No new PTN or family-wide evolutionary inference is asserted.

## Literature and experimental provenance

The 2021 membrane review (PMID34046426) and 2022 archaeal-lipid structural review
(PMID35976526) are read in full, assessed and mined into references/findings and
step evidence. The 2023 terpenome review (PMID36941124) is retained as broad
abstract-level context with its correction PMID37455337 explicitly recorded.
Its unread corrected figures provide no new evolutionary or activity claims.

[Peterhoff et al. 2014](https://onlinelibrary.wiley.com/doi/full/10.1111/mmi.12596)
compares enzymes using both purified-protein assays and a Bacillus complementation
host. The host supplies downstream lipid processing. Enzyme activity in that
host cannot establish native downstream capacity in the protein's source species.
The official PDB mappings independently distinguish bacterial 4NAE/4NAF, 3W01/3W02
and 4JEJ from archaeal 4MM1.

The 2025 reconstitution (PMID40626731) substitutes a C20 geranylgeranyl analogue
for the C35 native donor. Native genetic requirement and earlier lipid chemistry
support the C35 pathway. Reconstitution neither resolves endogenous G1P supply
nor demonstrates membrane crossing. An unresolved donor regioisomer preference
must not be converted into a substrate-specific claim.

## Limits that remain after curation

Native EgsA-to-PcrB flux, the native phosphatase, carrier membrane crossing,
donor regioisomer preference, and complete pathways beyond the characterized
Bacillus system remain open. These are biological limits, not missing identifiers
to be guessed. The 2012 dimerization paper has an indexed erratum whose contents
remain to be verified before adding detailed residue/interface claims.

Gene review consultations, focused checks and a complete repository validation
must finish before publication. Source caches and original annotation payloads
are preserved by the baseline and seed receipts.