Overview
OpenAntigens is generated as a local, versioned computational snapshot. Completed reports combine target identity, topology, extracellular or membrane-protein design scope, structure confidence, experimental structure precedent, domain/family context, ortholog mappings, BLAST searches, disease associations when enabled, and precomputed construct candidates.
Reports provide computational evidence for antigen construct design and antibody-discovery planning. Final construct choice still requires biological review, expression testing, and experimental validation.
Target universe and resolution
OpenAntigens Mouse contains unique mouse orthologs resolved from the OpenAntigens human target universe. Targets without a resolved mouse record are omitted, and duplicate mouse records are included once. Each included target is analyzed against reviewed mouse UniProt data; the originating human target remains in report metadata.
Mouse records are resolved through reviewed mouse UniProt entries. The build records skipped human targets, including missing mouse orthologs and duplicate mouse targets, in a separate manifest.
Topology and construct scope
Topology controls which design track is used. Secreted proteins use the mature extracellular protein as the soluble design scope. GPI-anchored proteins and single-pass proteins emphasize extracellular regions, with boundaries inferred from curated topology, signal peptide, propeptide, and transmembrane annotations.
Multipass proteins are treated separately. When the largest extracellular region exceeds 80 amino acids, OpenAntigens treats the protein as a mixed case with both soluble extracellular-region constructs and full-length membrane-expression context. With shorter extracellular loops, the report emphasizes membrane-protein expression and full-length context.
Sequence canonicality
OpenAntigens requires the report sequence, UniProt sequence, and AlphaFold model sequence to represent the same canonical isoform. Mismatching AlphaFold models are excluded from structure-dependent metrics because residue-number mismatches corrupt construct boundaries, pLDDT coloring, and sequence-to-structure selection.
AlphaFold DB models are used only when they match the canonical report sequence.
When a target lacks a matching AlphaFold DB model, non-structure-dependent annotations remain available; structure-derived segmentation and interactive structure features are limited.
AlphaFold confidence
pLDDT is used as local residue-confidence evidence. High pLDDT supports local fold confidence, while low pLDDT is consistent with flexible linkers, disordered tails, or uncertain loops. PAE is used as regional confidence evidence: low internal PAE supports a coherent structural unit, while high PAE between neighboring blocks indicates uncertain relative orientation.
Construct diagnostics summarize mean pLDDT, structured fraction, mean intrablock PAE, interblock PAE, local boundary PAE, separation, and linker pLDDT when available. These metrics are evidence features, not absolute pass/fail criteria.
Residue topology and accessibility
Residues are classified by topology first, using UniProt topology features, transmembrane segments, signal peptides, propeptides, mature-chain annotations, and the input topology track. The reported location categories are extracellular, membrane, intracellular, or unknown. For multipass proteins, extracellular termini and loops are treated as extracellular regions inside a membrane-protein model.
Surface accessibility is computed from the canonical AlphaFold structure with FreeSASA using the Lee-Richards algorithm. The default probe radius is 1.4 angstroms, the default slice count is 20, and residues are called solvent-accessible when relative SASA is at least 0.20.
Extracellular accessible residues
Antibody recognition depends most directly on residues that are both extracellular and accessible. OpenAntigens therefore tracks an extracellular accessible residue set and uses it in ortholog identity, paralog/family identity, BLAST summaries, and alignment highlighting.
PAE-block SASA is used because AlphaFold sometimes packs flexible domains or disordered segments against each other even when PAE indicates their relative placement is uncertain. Recomputing accessibility inside the PAE-coherent block reduces false buried calls for residues that may be exposed in solution or on cells.
The extracellular accessible identity reported in homolog, family, and BLAST sections is calculated only over aligned primary-target residues that are both extracellular and classified as exposed or conditional.
Construct generation
The analysis engine scores construct candidates, removes exact duplicate boundaries across calculated and annotated candidate sources (PDB-backed constructs are kept individually as evidence), filters soluble candidates outside the inferred design scope or below the minimum length, and the portal displays the surviving classes in a fixed review order.
The dedicated Pre-generated Constructs page expands this section with construct classes, diagnostics, card fields, homolog transfer, PTM-aware review, and practical selection guidance.
The minimum construct size defaults to 50 amino acids. Soluble constructs outside the inferred design scope are excluded. Exact duplicate boundaries from different calculated or annotated sources are collapsed during construct deduplication; PDB-backed constructs are exempt, so every deposited structure is retained as evidence.
Strict and lenient calculated constructs come from two AlphaFold segmentation passes: a conservative strict pass that isolates compact, domain-like units, and a permissive lenient pass that preserves larger, sometimes multi-domain regions. Lenient constructs are often larger and more epitope-preserving; strict constructs are smaller and more single-domain-like. The exact pLDDT thresholds, PAE split criteria, and boundary scoring are documented on the Pre-generated Constructs page.
Domain annotations
UniProt feature annotations and InterPro records are merged into the residue annotation map. Domain annotations must lie inside the relevant design scope to be used as construct recommendations. Large whole-protein signatures that extend beyond the mature secreted or extracellular region are retained as descriptive context when appropriate. They are excluded from soluble construct recommendations.
InterPro is treated as a higher-level curated domain/family source. Pfam entries support domain annotation. Pfam-only family IDs are excluded as canonical family identifiers for precomputed family matrices.
Family and paralog context
Canonical family assignment prefers InterPro family annotations when they are specific and informative. HGNC and Ensembl-derived paralog references are used to recover biologically relevant families when InterPro is incomplete or too broad. If multiple families are available, smaller and more specific families are preferred over large generic families.
Large families can be skipped from detailed matrix rendering to keep pages readable. Directional identity matrices are used because X-to-Y identity and Y-to-X identity can differ when one protein is a subregion or short paralog relative to another.
Orthologs and homology
The mouse portal keeps the originating human target in report metadata and maps additional homolog regions when suitable reference sequences are available.
Construct-level homolog sequences are inferred by aligning the mouse design scope to available homolog sequences, projecting the selected mouse boundaries through that alignment, and extracting the corresponding homolog region.
Experimental structures
PDB/RCSB evidence is mapped back to UniProt residue numbering where possible. Experimental construct boundaries that overlap the design scope are summarized and can become PDB-backed construct suggestions. PDB entries fully outside the mature secreted, extracellular, or relevant membrane-protein scope are ignored for construct recommendation.
PDB evidence is treated as design precedent. Exact construct choice still depends on the current antibody-discovery context.
BLAST sequence similarity and cross-reactivity context
Local blastp searches provide sequence similarity context for reviewing possible cross-reactivity. Mouse target sequences are searched against the configured local human, mouse, and cynomolgus monkey protein databases.
Self hits are filtered by accession and entry name. BLAST results are ranked sequence similarity signals, not measurements of antibody cross-reactivity. No universal threshold is applied because acceptable risk depends on antigen class, screening goal, and downstream assay design.
PTMs and processing features
OpenAntigens extracts curated UniProt PTM-like features including glycosylation, modified residues, lipidation, disulfide bonds, cross-links, initiator methionine processing, signal peptides, propeptides, mature chains, peptides, and cleavage-related site annotations.
Every target page lists these annotations in a dedicated PTM section. Construct cards list which PTMs overlap each construct, and the Interactive Construct Builder updates the included PTMs in real time as the selected residue window changes. Processing and cleavage annotations raise boundary-review warnings because cutting through or including these sites unintentionally can affect expression and antigen quality.
Cysteine analysis
Cysteines are reported as paired or unpaired when curated disulfide evidence or inferred pairing context is available. Unpaired cysteines inside a construct trigger warnings, especially when they appear accessible or are associated with unwanted disulfides, aggregation, or expression heterogeneity.
The Interactive Construct Builder can optionally apply Cys-to-Ser edits to selected unpaired cysteines and updates copied TSV/FASTA sequences in real time.
Furin-site scanning
Basic furin-like cleavage motifs are scanned across the design region and constructs. The pipeline reports suggested edits as annotations and leaves application to manual review because cleavage risk is context-dependent and some basic motifs may be biologically important.
Ligand and interaction regions
Curated UniProt features, PDB mappings, and annotation text are used to report ligand-binding or interaction-related regions when available. These annotations help users avoid trimming known functional surfaces that may be needed for conformational antibody discovery.
Assembly and partner requirements
Interaction and complex information is summarized separately from obligatory-partner classification. Known interactions provide context; obligatory-partner warnings require stronger curated complex evidence.
Obligatory-partner warnings are anchored to curated Complex Portal records when that module is enabled. UniProt SUBUNIT comments are retained as interaction and assembly context without independently creating obligatory-partner warnings.
This conservative behavior avoids false mandatory-partner calls that would incorrectly discourage soluble antigen designs. Reports can list many known interactions without an obligatory requirement when curated evidence falls short of the stronger mandatory-partner criteria.
Disease, literature, and antibody-resource context
The mouse portal does not display Open Targets disease associations or disease-search controls.
PubTator3 is queried for approximate gene-linked PubMed hit counts. These counts support triage and literature awareness. Target validity requires separate curation. CiteAb links are provided as external antibody-resource shortcuts; CiteAb catalog content remains external.
Static assets and visualization
When assets are generated, each construct can include structure images, pLDDT plots, and PAE plots with the construct region highlighted. Structure renderings use AlphaFold models colored by pLDDT and emphasize cysteines when possible. The portal also includes an Interactive Construct Builder that links sequence selection, structure highlighting, pLDDT, PAE, warnings, and copy-ready sequence outputs.
Static image assets support report review. Interactive widgets support boundary tuning and exploratory design.
Update cadence and limitations
OpenAntigens is released as a static, versioned snapshot. Each release records the build date, target universe, and OpenAntigens version when available. Future releases may incorporate updated annotations.
Because each release is static, source databases may have changed after the displayed release date. Cite the release date and downloaded artifact names when using OpenAntigens data in downstream analyses.
