LY96 (MD-2) review notes

UniProt Q9Y6Y9, lymphocyte antigen 96 / myeloid differentiation protein 2. 160 aa,
signal peptide 1-18, ML (MD-2-related lipid-recognition) domain, no transmembrane
segment. Isoform 2 lacks residues 38-68.

Deep research status

LY96-deep-research-asta.md (Asta corpus retrieval) returned 20 generic
bioinformatics/database papers unrelated to MD-2 and was not used. The review was
built from the UniProt record, the cached GOA publications (all 14 cached; several
abstract-only) and the cached Reactome entries.

Deep-research cross-check (2026-09-30)

Compared the completed review against LY96-deep-research-falcon.md (sources are
mostly 2021-2025 reviews, a dissertation and Fu et al. 2025 cryo-EM).

• Agreement: MD-2 is the non-enzymatic LPS/lipid A-binding coreceptor of TLR4; TLR4
  alone lacks the recognition pocket; beta-cup fold with a hydrophobic cavity; binds the
  TLR4 ectodomain via a surface including the C95-C105 region; agonist-bound MD-2 (F126
  loop, one exposed acyl chain, phosphates) drives the 2:2:2 TLR4-MD-2-LPS complex;
  lipid IVa is a species-dependent antagonist in human; soluble/secreted MD-2 exists;
  CD14-dependent endocytosis feeds the TRAM/TRIF branch. All of these are already
  covered by the MF/CC/BP decisions, the description and core_functions.
• Additions (not used to change annotations): Asn26/Asn114 glycosylation requirement
  (dissertation source, not verified); angiotensin II as a direct MD-2 ligand (review
  source, not verified) -- added to the existing suggested question on non-LPS ligands;
  VEO-IBD LY96 p.T116del variant, DCM biomarker data, drug/antagonist catalogue, and the
  zebrafish ly96 ortholog -- context only, no GO consequence.
• MD-2 supports TLR4 maturation and surface delivery: verified as a statement in a
  cached primary paper PMID:11593030.
  No GOA row captures this and no NEW term was proposed (it would be a regulation/
  chaperone-type term whose actor question is unresolved); added as a suggested question.
• Conflicts: none. The report's framing ("principal ligand-binding component",
  "TLR4 supplies the transmembrane and intracellular signaling machinery") matches the
  review's choice of coreceptor activity (enables) plus contributes_to GO:0001875 in
  core_functions, and is consistent with TLR4 (enables GO:0001875, core) and CD14
  (GO:0001875 -> molecular carrier activity).
• Changes: no annotation actions changed. Added the Falcon report to references;
  added PMID:11593030 (surface-expression quote) and the Falcon report as extra
  support/reason text on the GO:0031666 IBA row (action stays KEEP_AS_NON_CORE); two
  suggested-question edits.

Biology, with provenance

• Discovery: MD-2 is needed for TLR4 to respond to LPS.
  PMID:10359581
  PMID:10359581
  MD-2 reaches the surface only with TLR4:
  PMID:10359581
• Secreted protein, disulfide-linked oligomers, soluble MD-2 is active:
  PMID:11593030
  PMID:11593030
• Two separable surfaces: TLR4-binding (C95-C105 region) and LPS-responsiveness (basic/aromatic region).
  PMID:14607928
  PMID:14607928
• LPS is crosslinked to MD-2 and TLR4 in the CD14-containing complex:
  PMID:11274165
• Structures: MD-2 binds the concave face of TLR4 ectodomain; ligand sits in MD-2's hydrophobic pocket.
  PMID:17803912
  PMID:17803912
  PMID:19252480
  PMID:22532668
  PMID:22532668
  PMID:22532668
• Not involved in LTA sensing via TLR2 (at least for the LTA preparations tested):
  PMID:12594207
• Other ligands of the MD-2 pocket: morphine
  PMID:22474354;
  heme (Reactome R-HSA-9707594); HMGB1 binds the TLR4/MD-2 complex PMID:20547845.
• Der p 2 (house dust mite allergen) is an MD-2 structural/functional mimic and co-IPs with MD-2
  PMID:19060881.

Which receptor MF belongs to MD-2 vs TLR4

• The recognition step (binding lipid A) is performed by MD-2's pocket; TLR4 alone does not
  bind LPS PMID:22532668. The one exposed acyl chain and the phosphates contact TLR4 and
  drive the 2:2:2 dimerisation PMID:19252480.
• Transmembrane transmission and TIR-domain signalling belong to TLR4 alone; MD-2 has no
  transmembrane or cytoplasmic part.
• So: MD-2 directly enables LPS binding (GO:0001530) and coreceptor activity (GO:0015026 —
  "combining with an extracellular messenger, and in cooperation with a nearby primary
  receptor, initiating a change in cell activity" fits MD-2 exactly), and contributes to
  GO:0001875 LPS immune receptor activity, which is a property of the TLR4-MD-2 heterodimer.
  The existing enables GO:0001875 rows (IBA + IDA PMID:19252480) are accepted: MD-2 is the
  ligand-binding subunit of a two-chain receptor and does the recognition half of the work;
  in core_functions it is recorded as contributes_to.
• Process: detection of lipopolysaccharide (GO:0032497) is a direct participation — MD-2
  performs the detection step. TLR4 signalling pathway: MD-2 is part of the receptor that
  initiates it.

Annotation decisions (summary)

• protein binding IPI rows with TLR4 partner -> MODIFY to GO:0035662 Toll-like receptor 4 binding.
• protein binding with Der p 2, LCN2, MESD -> REMOVE (uninformative; xeno allergen mimic / HuRI Y2H).
• cellular defense response (TAS) -> MODIFY to innate immune response.
• cell surface receptor signaling pathway (TAS) -> MODIFY to toll-like receptor 4 signaling pathway.
• positive regulation of LPS-mediated signaling (IBA) -> KEEP_AS_NON_CORE: MD-2 is a receptor
  subunit rather than a regulator, though soluble MD-2 does set cell responsiveness.
• positive regulation of TNF production -> KEEP_AS_NON_CORE (downstream output).
• endosome membrane (Reactome) -> KEEP_AS_NON_CORE (TLR4-MD-2 internalised for TRAM/TRIF branch).
• No NEW annotations.