amiB2 Resolved · high 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.

In the literature (TB corpus sweep) 1 publication

1 TB publication mentions this gene. 1 publication(s) discuss this gene (0 in a M. tuberculosis context, 1 in other mycobacteria — 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.

CRISPRi vulnerability

Vulnerability index 0.76 (95% CI -1.02 to 3.43). 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 Mycobrowser is no longer maintained; its EC numbers predate recent nomenclature revisions, so a class change usually reflects re-numbering, not a conflict.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)

UniProt still lists this protein as Putative amidase AmiB2; the revised annotation above is ahead of the current UniProt record.

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) Bacteria

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)

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.

Enzyme activity (activity-based protein profiling) active serine hydrolase confirms atlas call high-confidence target

Directly labelled by a fluorophosphonate activity-based probe in live M. tuberculosis: biochemical proof that this protein is a catalytically active serine hydrolase, not merely a predicted fold. This experimentally corroborates the atlas assignment (Amidase. Pfam: Amidase (PF01425.27).), which was derived independently from structure and orthology.

experimental (activity-based) confirmation of the atlas serine-hydrolase family assignment; proves the enzyme is catalytically active in live M. tuberculosis. It never changes the verdict here. Source: Li M, Patel HV, ..., Canaan S, Aldridge BB et al., Cell Chem Biol 2021 (PMC8964833; doi:10.1016/j.chembiol.2021.09.002); competitive activity-based protein profiling of FP-reactive serine hydrolases.

Mutant phenotypes (conditional Tn-seq, MtbTnDB)

Conditional fitness of transposon-disruption mutants across 1 significant condition(s) (|log2FC|≥1, q≤0.05), from the standardized MtbTnDB compendium. A negative log2FC means the mutant is depleted — the gene contributes to fitness in that condition. An in-vivo defect for a "hypothetical" is strong evidence it matters for infection, even without a known molecular function. Disruption (Tn insertion), not a clean deletion; genetic-interaction screens excluded.

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.

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)

Structural search (AlphaFold DB model, Foldseek vs PDB — genome-wide) pLDDT 97.0

Foldseek search of the AlphaFold DB model (mean pLDDT 97.0, gated at 70) against the PDB — a genome-wide extension of the ESMFold dark-gene search that also covers proteins beyond the single-sequence length limit. Confident structural neighbours (E < 0.01) shown.

Genomic context (neighbours & predicted operon)

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).

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: gatB (aspartyl/glutamyl-tRNA(Asn/Gln) amidotransferase subunit B), high confidence from genomic context alone (score 968 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

• Legacy H37Rv annotation: amidase AmiB
• MTBC0 PGAP product: amidase
• Pfam (hmmscan --cut_ga): Amidase PF01425.27 (E=2e-100)
• (auto-curated by rules from PGAP + Pfam + Foldseek; not hand-reviewed)

Ancestral MTBC0 protein sequence

>mtbc0_001352|Rv1263|amiB2
MDPTDLAFAGAAAQARMLADGALTAPMLLEVYLQRIERLDSHLRAYRVVQFDRARAEAEAAQQRLDAGERLPLLGVPIAIKDDVDIAGEVTTYGSAGHGPAATSDAEVVRRLRAAGAVIIGKTNVPELMIMPFTESLAFGATRNPWCLNRTPGGSSGGSAAAVAAGLAPVALGSDGGGSIRIPCTWCGLFGLKPQRDRISLEPHDGAWQGLSVNGPIARSVMDAALLLDATTTVPGPEGEFVAAAARQPGRLRIALSTRVPTPLPVRCGKQELAAVHQAGALLRDLGHDVVVRDPDYPASTYANYLPRFFRGISDDADAQAHPDRLEARTRAIARLGSFFSDRRMAALRAAEVVLSSRIQSIFDDVDVVVTPGAATGPSRIGAYQRRGAVSTLLLVVQRVPYFQVWNLTGQPAAVVPWDFDGDGLPMSVQLVGRPYDEATLLALAAQIESARPWAHRRPSVS