PPE4 Family assigned · medium auto-curated

Annotation: from legacy to revised

Auto-curated: this verdict and function were generated by rules from PGAP + Pfam + Foldseek and have not been hand-reviewed.

Annotated on the H37Rv protein: this gene has no 1:1 ancestral MTBC0 anchor (PE/PPE, paralogue, IS element, or otherwise unanchored CDS).

In the literature (TB corpus sweep) 6 publications

6 TB publications mention this gene. 6 publication(s) discuss this gene (6 in a M. tuberculosis context, 2 in other mycobacteria — M. leprae (1), M. smegmatis (1)).

This layer CITES the literature and adds context; it does not change the verdict or the function stated elsewhere in this fiche. This distinguishes a gene that is dark because nobody has looked from one that is dark despite having been studied. Source: PubMed (whole): H37Rv locus tag + GENE NAME + ortholog identifiers (Mb…, MMAR_…, MSMEG_…, ML…, MAB_…), under a mycobacterial context filter; hits verified against the abstract text. Species-context counts distinguish M. tuberculosis literature from literature on other mycobacteria. phase76/phase77, 2026-07-13.

Intrinsic disorder (sequence + structure) highly disordered

carries a substantial disordered region (272/513 residues); disorder is a property, not a function

A property (biophysics), not a function. No LLPS/condensate claim is made from disorder alone. Verdict unchanged. Source: metapredict v3 (Emenecker/Holehouse) per-residue disorder + AlphaFold mean pLDDT (annotation_mtbc P16.13).

Conditional expression context (iModulons)

Member of 2 independently-modulated gene set(s): IdeR (ideR), Zur (zur).

iModulon membership (independently-modulated gene sets from a 647-sample RNA-seq compendium): the conditional co-expression context. Co-expression is a regulatory context, NOT a molecular function. Source: iModulonDB / modulome_mtb (Yoo 2022).

CRISPRi vulnerability

Vulnerability index -6.32 (95% CI -6.87 to -5.78). A more negative index = more vulnerable to knockdown (better drug-target quality); indicative threshold VI ≤ -6 = highly vulnerable.

Quantitative CRISPRi knockdown, graded (finer than binary Tn-seq essentiality). Source: CRISPRi vulnerability index (Bosch 2021, pebble.rockefeller.edu).

Legacy record & comparison (Mycobrowser)

The legacy Mycobrowser record is shown for verification. Mycobrowser is no longer maintained; its EC numbers predate recent nomenclature revisions, so a class change usually reflects re-numbering, not a conflict.

Orthologues (reciprocal best hits across mycobacteria)

Reciprocal-best-hit orthologues (DIAMOND) against the Mycobrowser reference proteomes. A missing species is informative: e.g. a gene absent from M. leprae was likely lost in its reductive genome evolution. Locus tags link to Mycobrowser.

Curated reference (UniProt)

Functional vocabulary (eggNOG-mapper, orthology transfer)

Orthology-based transfer (eggNOG 5.0.2, diamond). EC/KO/GO/CAZy are computed annotations, not manual curation; cross-check against the primary literature before treating a specific reaction as established.

Conservation & selection (intra-MTBC, 145 209 strains)

pN/pS from segregating SNPs (singletons removed) normalised by possible sites. Low pN/pS = purifying selection (a strong signal that a "hypothetical" is a real, constrained gene). A high pN/pS is ambiguous: relaxed constraint or positive selection (drug resistance, antigenic variation) inflate it; e.g. rpoB/katG/pncA score high here for resistance, not loss of function. A clonal disruption (one allele over a clade) suggests lineage pseudogenisation; a convergent one (many independent alleles) is typical of resistance loss-of-function.

Outgroup conservation (beyond the MTBC) Mycobacteriaceae

Two orthogonal outgroup signals. M. canettii (the immediate outgroup) gives a deep-divergence dN/dS (a low value confirms a constrained, real gene; shown as confident only at ≥8 substitutions, else flagged low-power). Genus-wide presence/absence (tblastn vs assembled non-MTBC genomes) places the gene on the ancient-core ↔ MTBC-specific axis: a gene absent even from the closest MTBAP relatives is a candidate MTBC-specific innovation (possible host-adaptation factor, to confirm by synteny). The phylostratum extends that axis outside the genus (tblastn vs 13 reference genomes spanning Mycobacteriaceae → Corynebacteriales → Actinomycetia → outside the phylum): it is the deepest clade in which a homolog is still detected, i.e. a proxy for gene age. Read it with the null model in mind: a shallow (young) stratum can also reflect homology-detection failure for short or fast-evolving ORFs, so it is a descriptive axis, not a proof of novelty.

Essentiality (transposon mutagenesis) essential

Genome-wide Himar1 transposon essentiality in H37Rv (DeJesus 2017). An essential call (ES/ESD/GD) is strong, independent evidence that a "hypothetical" locus encodes a functional, selectively required gene — orthogonal to intra-species conservation.

Chemical-genetic target & druggability (PROSPECT) hypomorph tool strain

This gene is part of the PROSPECT collection of TetON transcriptional-knockdown (hypomorph) strains of essential M. tuberculosis genes, built as a sensitised background for chemical-genetic mechanism-of-action deconvolution. Being in the panel means the gene is an essential / vulnerable target for which a validated knockdown tool strain exists.

Panel membership reflects essentiality/vulnerability and the availability of a genetic tool, not a specific molecular function; it never changes the verdict here. Source: Bond AN et al., Nat Commun 2025;16:9673 (doi:10.1038/s41467-025-64662-x); PROSPECT chemical-genetic platform.

Proteomics (mass spectrometry) detected

Detection by mass spectrometry is direct, experimental evidence that the protein product exists — orthogonal to sequence conservation and to Tn-seq essentiality, and especially decisive for a "hypothetical" locus. Reproducible detection across several independent datasets (PaxDb) makes the existence claim robust; the integrated abundance places the protein in the proteome's dynamic range.

Predicted localisation (DeepTMHMM + lipobox)

Transmembrane topology and signal peptide from DeepTMHMM (deep-learning reference predictor); lipoproteins from a (myco)bacterial lipobox motif. A sequence-based prediction of subcellular context.

Physico-chemical properties (computed, ProtParam)

Computed from the ancestral MTBC0 sequence with the ExPASy ProtParam method (Biopython). Descriptive biophysical context: a positive GRAVY flags a hydrophobic (often membrane) protein, a high instability index (>40) predicts a short in-vitro half-life, an extreme pI hints at compartment or binding partner.

Domains (Pfam, hmmscan --cut_ga)

Experimental structures (Protein Data Bank) 2 solved

Experimentally solved structures mapped from the UniProt accession via PDBe/SIFTS (2 total; up to 8 shown, ranked by sequence coverage then resolution). An experimental structure is direct proof of the folded product and the strongest structural evidence — superseding the predicted ESMFold/AlphaFold models below for any covered region.

Genomic context (neighbours & predicted operon) operon of 11

Neighbours from the H37Rv annotation (+ strand). The operon is predicted by co-directional intergenic distance (same strand, gaps ≤50 bp) — a transcription-unit hypothesis, not a mapped TSS. For a "hypothetical", co-transcription with a characterised operon is a concrete functional lead (complements the STRING neighborhood channel below).

Transcriptional regulation (signed TRN: ChIP-seq + TFOE)

Regulatory edges from the ISB signed transcriptional regulatory network (TF ChIP-seq binding, Minch 2015 + TF-overexpression response, Rustad 2014). An edge is regulatory evidence (binding and/or expression change), not necessarily direct. For a "hypothetical", membership in a known regulon (e.g. DosR dormancy, PhoP virulence) is a strong physiological-context lead.

Node colour = verdict, dashed = hypothetical; edge colour = evidence (green experimental, orange genomic-context, grey co-expression), width ∝ score. Click a partner to open its page; "Explore full network" to walk the graph.

Closest characterised functional partner: eccC3 (ESX-3 secretion system protein EccC3), high confidence from genomic context alone (score 958 excluding text-mining).

STRING combines evidence channels (neighborhood, fusion, cooccurrence, coexpression, experimental, database, text-mining) into a 0–1000 score. The ctx badge marks edges carried by the genomic-context channels (conserved neighborhood, fusion, phylogenetic co-occurrence), which are independent of orthology and structure and the strongest signal for an unknown gene. The exp badge marks an experimentally-supported partner (measured interaction, experimental/database channel ≥400) as opposed to a purely predicted one — but note that the M. tuberculosis experimental interactome is dominated by a noisy bacterial-two-hybrid screen, so a strong measured link that contradicts the operon/localisation context is likely a false positive. The no text-mining column recomputes the score from data alone, so a link that does not depend on the literature is visible. Association is a function hypothesis, not proof: corroborate with the operon context and the primary literature before assigning a function.

Evidence

• Annotation from H37Rv (no MTBC0 1:1 anchor; H37Rv protein used): PPE family protein PPE4
• Pfam (hmmscan --cut_ga): PPE PF00823.26 (E=1e-62), PPE-PPW PF18878.6 (E=9e-21)
• (auto-curated by rules from PGAP + Pfam + Foldseek; not hand-reviewed)

Ancestral MTBC0 protein sequence

>H37Rv|Rv0286|PPE4
MAAPIWMASPPEVHSALLSNGPGPGSLVAAATAWSQLSAEYASTAAELSGLLGAVPGWAWQGPSAEWYVAAHLPYVAWLTQASADAAGAAAQHEAAAAAYTTALAAMPTLAELAANHVIHTVLVATNFFGINTIPITLNEADYVRMWLQAAAVMGLYQAASGAALASAPRTVPAPTVMNPGGGAASTVGAVNPWQWLLALLQQLWNAYTGFYGWMLQLIWQFLQDPIGNSIKIIIAFLTNPIQALITYGPLLFALGYQIFFNLVGWPTWGMILSSPFLLPAGLGLGLAAIAFLPIVLAPAVIPPASTPLAAAAVAAGSVWPAVSMAVTGAGTAGAATPAAGAAPSAGAAPAPAAPATASFAYAVGGSGDWGPSLGPTVGGRGGIKAPAATVPAAAAAAATRGQSRARRRRRSELRDYGDEFLDMDSDSGFGPSTGDHGAQASERGAGTLGFAGTATKERRVRAVGLTALAGDEFGNGPRMPMVPGTWEQGSNEPEAPDGSGRGGGDGLPHDSK