Affinage

MCFD2

Multiple coagulation factor deficiency protein 2 · UniProt Q8NI22

Audit flag: ungrounded claim
Length
146 aa
Mass
16.4 kDa
Annotated
2026-06-10
20 papers in source corpus 13 papers cited in narrative 12 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 7/7 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

MCFD2 is a soluble, calcium-binding cargo-recognition subunit of the early secretory pathway that pairs with the transmembrane lectin LMAN1 (ERGIC-53) to mediate efficient ER-to-Golgi transport of selected secretory glycoproteins, most notably coagulation factors V and VIII (PMID:15886209, PMID:17010120). MCFD2 and LMAN1 form a 1:1, calcium-dependent complex; MCFD2 is retained in the ER through this interaction and is secreted when LMAN1 is absent (PMID:15886209, PMID:17010120). Structurally, MCFD2 is intrinsically disordered in the apo state and folds upon binding Ca2+ to its two C-terminal EF-hand motifs, which are necessary and sufficient for LMAN1 binding, while the N-terminal region is dispensable (PMID:18590741, PMID:20007547). The EF-hand region also engages cargo (FV/FVIII) through binding sites separable from the LMAN1 interface, such that mutations disrupting Ca2+-induced folding and LMAN1 binding still retain cargo binding (PMID:20007547). On the receptor side, LMAN1 contacts MCFD2 via the N-terminal β-sheet of its carbohydrate-recognition domain, distinct from its Ca2+/sugar-binding cargo and mannose sites, and must oligomerize to function as a cargo receptor (PMID:20817851, PMID:20138881). The complex acts with cargo selectivity—required for FV/FVIII but dispensable for cathepsin binding—and also promotes ER exit of α1-antitrypsin, where LMAN1's lectin-glycan interaction is critical (PMID:17010120, PMID:29735583, PMID:35822856). Within the complex MCFD2 carries out cargo binding and transport while LMAN1 primarily serves as a shuttling carrier, and disease-causing MCFD2 missense mutations (e.g., I136T) reduce complex affinity by orders of magnitude and cause combined factor V and factor VIII deficiency (PMID:16304051, PMID:18056485, PMID:36490287).

Mechanistic history

Synthesis pass · year-by-year structured walk · 11 steps
  1. 2005 High

    Established that MCFD2 physically partners with LMAN1 in a defined stoichiometry and directly contacts coagulation factor VIII, defining the molecular basis of the cargo receptor complex.

    Evidence Cross-linking immunoprecipitation, reciprocal Co-IP, and stoichiometry analysis in the early secretory pathway

    PMID:15886209

    Open questions at the time
    • Did not resolve which structural elements of MCFD2 mediate each interaction
    • Cargo binding site on MCFD2 versus LMAN1 not yet distinguished
  2. 2005 Medium

    Showed that disease-associated MCFD2 mutations need not fully abolish complex formation to cause clinically significant FV/FVIII deficiency, refining the genotype-phenotype relationship.

    Evidence Co-IP and Western blot of patient-derived lymphoblasts carrying the I136T mutation

    PMID:16304051

    Open questions at the time
    • Quantitative threshold of residual complex for secretion not defined
    • Single patient mutation, single lab
  3. 2006 High

    Defined cargo selectivity by demonstrating MCFD2 is specifically required for FV/FVIII but dispensable for other ERGIC-53 glycoprotein cargo, and that ERGIC-53 retains MCFD2 in the ER.

    Evidence siRNA knockdown and YFP fragment complementation assay for in vivo cargo binding

    PMID:17010120

    Open questions at the time
    • Molecular determinants of cargo selectivity unresolved
    • Did not address non-coagulation cargo such as AAT
  4. 2007 High

    Identified the C-terminus and calcium dependence as structural prerequisites for the MCFD2–ERGIC-53 interaction and quantified the affinity loss in disease mutants.

    Evidence Biochemical binding assays, C-terminal deletion analysis, surface plasmon resonance, and flow cytometry; calcium-dependence below 0.2 mM measured

    PMID:17971482 PMID:18056485

    Open questions at the time
    • Structural basis of disorder-to-order transition not yet visualized
    • How MCFD2 enhances ERGIC-53 sugar binding mechanistically unclear
  5. 2008 High

    Explained the calcium dependence at atomic resolution: MCFD2 is disordered without Ca2+ and folds upon binding to its EF-hands, while disease mutants fail to fold even with calcium.

    Evidence Solution NMR structure determination with circular dichroism of mutant variants

    PMID:18590741

    Open questions at the time
    • Structure of the bound complex with LMAN1 not yet determined
    • Cargo-bound conformation not resolved
  6. 2009 High

    Mapped the binding architecture, showing the C-terminal EF-hands are necessary and sufficient for LMAN1 binding and that cargo binding occurs through separate sites independent of Ca2+-induced folding.

    Evidence Deletion mutagenesis, Co-IP, and CD spectroscopy

    PMID:20007547

    Open questions at the time
    • Precise cargo-contacting residues on MCFD2 not identified
    • How a partially folded mutant retains cargo binding mechanistically unclear
  7. 2010 High

    Defined the receptor side structurally: LMAN1 binds MCFD2 via its CRD β-sheet, separate from its sugar/cargo sites, and requires oligomerization for cargo receptor function.

    Evidence Crystal structure of LMAN1-CRD/MCFD2 complex with site-directed mutagenesis and Co-IP

    PMID:20138881 PMID:20817851

    Open questions at the time
    • Full-length complex with cargo bound not crystallized
    • Stoichiometry of cargo loading per oligomer unknown
  8. 2011 High

    Genetic dissection in mice revealed non-identical roles of LMAN1 and MCFD2 and extended the receptor's cargo repertoire to α1-antitrypsin.

    Evidence LMAN1 and MCFD2 single and double knockout mice with plasma protein measurements and hepatocyte ER fractionation

    PMID:29735583

    Open questions at the time
    • Identity of the alternative FV/FVIII secretion pathway not determined
    • Mechanism distinguishing single- versus double-KO phenotypes unclear
  9. 2020 Medium

    Showed MCFD2 possesses conformational plasticity that LMAN1-CRD lacks, providing a structural rationale for accommodating diverse polypeptide cargo.

    Evidence High-resolution X-ray crystallography of multiple ERGIC-53-CRD/MCFD2 crystal forms

    PMID:32356523

    Open questions at the time
    • Functional role of plasticity inferred from structure only
    • No cargo-bound structure to confirm accommodation
  10. 2022 High

    Established the LMAN1-MCFD2 complex as a lectin-dependent cargo receptor for α1-antitrypsin, with LMAN1 binding AAT independently of MCFD2 via glycan recognition.

    Evidence CRISPR knockout cell lines, Co-IP, secretion/chase assays, and AAT glycosylation mutants including the Z variant

    PMID:35822856

    Open questions at the time
    • Why AAT depends on glycan binding while FV/FVIII can bypass it not reconciled
    • Quantitative contribution of MCFD2 to AAT transport not isolated
  11. 2023 Medium

    Reassigned functional division of labor: MCFD2 alone can drive FV/FVIII cargo binding and transport, with LMAN1 acting mainly as a shuttling carrier and its lectin activity dispensable for these cargoes.

    Evidence Overexpression rescue assays in LMAN1/MCFD2-deficient cell lines with functional secretion readouts

    PMID:36490287

    Open questions at the time
    • Apparent contradiction with glycan-dependent AAT transport unresolved
    • Single lab; mechanism of MCFD2-driven shuttling without LMAN1 lectin function unclear

Open questions

Synthesis pass · forward-looking unresolved questions
  • The identity of the alternative, LMAN1/MCFD2-independent secretion pathway for FV/FVIII and the structural basis for cargo loading onto the assembled receptor remain unresolved.
  • No cargo-bound structure of the full complex
  • Alternative secretion route uncharacterized
  • Reconciliation of glycan-dependent versus glycan-independent cargo transport pending

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0038024 cargo receptor activity 4 GO:0060090 molecular adaptor activity 3
Localization
GO:0005783 endoplasmic reticulum 3 GO:0005794 Golgi apparatus 2
Pathway
R-HSA-5653656 Vesicle-mediated transport 3 R-HSA-9609507 Protein localization 3 R-HSA-392499 Metabolism of proteins 2
Partners
LMAN1
Complex memberships
LMAN1-MCFD2 cargo receptor complex

Evidence

Reading pass · 12 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2005 LMAN1 and MCFD2 form a 1:1 stoichiometric complex in the early secretory pathway, and both proteins interact with coagulation factor VIII (at the B domain) via calcium-dependent protein-protein interactions, independent of factor VIII glycosylation state. MCFD2 is retained in the ER through its interaction with LMAN1, and MCFD2 interaction with FVIII is independent of LMAN1-MCFD2 complex formation. Cross-linking immunoprecipitation, co-immunoprecipitation, Western blot, stoichiometry analysis The Journal of biological chemistry High 15886209
2005 Missense mutations in MCFD2 (e.g., I136T) result in a low but detectable level of residual LMAN1-MCFD2 complex, suggesting complete loss of the complex is not required for clinically significant reduction in FV and FVIII secretion. Immunoprecipitation and Western blot of patient-derived lymphoblasts Blood Medium 16304051
2006 MCFD2 is dispensable for binding of cathepsin Z and cathepsin C to ERGIC-53, demonstrating cargo selectivity: MCFD2 is specifically required for recruitment of coagulation factors V and VIII but not for glycoprotein cargo in general. In the absence of ERGIC-53, MCFD2 is secreted rather than retained. siRNA knockdown, yellow fluorescent protein fragment complementation assay (in vivo cargo binding) Traffic (Copenhagen, Denmark) High 17010120
2007 Deletion of the C-terminal 3 residues (ΔS-L-Q) of MCFD2 impairs binding to ERGIC-53 due to modification of the 3D structure of MCFD2, establishing that the C-terminus is structurally critical for the ERGIC-53/MCFD2 interaction. Biochemical binding assay, structural analysis of mutant MCFD2 Blood Medium 17971482
2007 MCFD2 interaction with ERGIC-53 enhances the sugar-binding ability of ERGIC-53 (specifically for high-mannose oligosaccharides, especially M8B). F5F8D patient MCFD2 missense mutants show 3–4 orders of magnitude lower affinity for ERGIC-53 by surface plasmon resonance. The MCFD2-ERGIC-53 interaction is calcium-dependent, becoming significantly weaker below 0.2 mM calcium. Flow cytometry binding assay, surface plasmon resonance, endo H treatment Blood High 18056485
2008 The solution structure of human MCFD2 determined by NMR shows the protein is disordered in the apo (calcium-free) state and folds upon binding Ca2+ to its two C-terminal EF-hand motifs. Disease-causing missense mutants are predominantly disordered even in the presence of calcium, explaining the calcium-dependence of the MCFD2-ERGIC-53 interaction. Solution NMR structure determination, circular dichroism of mutant variants Journal of molecular biology High 18590741
2009 The C-terminal EF-hand domains of MCFD2 are both necessary and sufficient for interaction with LMAN1; the N-terminal non-EF-hand region is dispensable for LMAN1 binding. The EF-hand domains also mediate interaction with FV and FVIII, but through separate binding sites: mutations abolishing LMAN1 binding (and disrupting tertiary structure) still retain FV/FVIII binding, indicating FV/FVIII interaction is independent of Ca2+-induced folding. Deletion mutagenesis, co-immunoprecipitation, circular dichroism spectroscopy Blood High 20007547
2010 Crystal structure of the LMAN1-CRD/MCFD2 complex reveals that LMAN1 interacts with MCFD2 through its N-terminal β-sheet of the CRD; mutations in the first β-sheet abolish MCFD2 binding without affecting mannose binding. Mutations in the Ca2+- and sugar-binding sites of the CRD disrupt FV/FVIII interaction without affecting MCFD2 binding, demonstrating distinct, separable binding sites for MCFD2 and cargo (FV/FVIII) on LMAN1. Monomeric LMAN1 mutants are defective in ER exit and unable to interact with MCFD2, showing oligomerization is required for cargo receptor function. Crystal structure, site-directed mutagenesis, co-immunoprecipitation Blood / FEBS letters High 20138881 20817851
2011 MCFD2-deficient mice have lower plasma FV and FVIII levels than LMAN1-deficient mice. Doubly deficient (LMAN1/MCFD2) mice show FV/FVIII levels matching LMAN1-deficient mice, suggesting an alternative secretion pathway exists. Both LMAN1 and MCFD2 are required for efficient ER exit of α1-antitrypsin (AAT), as demonstrated by reduced plasma AAT and accumulation in hepatocyte ER in singly and doubly deficient mice. Gene targeting (knockout mice), plasma protein level measurement, hepatocyte ER fractionation Blood advances High 29735583
2020 Crystallographic snapshots of ERGIC-53-CRD/MCFD2 complexes reveal that MCFD2 exhibits significant conformational plasticity whereas ERGIC-53-CRD does not, suggesting that MCFD2's structural flexibility is relevant to its ability to accommodate various polypeptide cargo ligands. X-ray crystallography (multiple crystal forms, 1.60 Å resolution) Acta crystallographica. Section F, Structural biology communications Medium 32356523
2022 LMAN1 and MCFD2 function as a cargo receptor complex for ER-to-Golgi transport of α1-antitrypsin (AAT). LMAN1 or MCFD2 KO cells show reduced AAT secretion and elevated intracellular AAT due to delayed ER-to-Golgi transport. AAT interaction with LMAN1 is independent of MCFD2 (by Co-IP in MCFD2 KO cells). Elimination of the second N-glycosylation site of AAT abolished LMAN1-dependent secretion, indicating lectin-glycan interaction is critical. Secretion of AAT Z-variant (monomers and polymers) is also LMAN1-dependent. CRISPR/KO cell lines, Co-immunoprecipitation, secretion/chase assays, glycosylation mutant analysis The Biochemical journal High 35822856
2023 Overexpression of either wild-type or mutant MCFD2 alone is sufficient to rescue FV/FVIII secretion defects in LMAN1-deficient cells, suggesting that MCFD2 carries out cargo binding and transport while LMAN1 primarily serves as a shuttling carrier for MCFD2. N-glycan binding by LMAN1 is not essential for FV/FVIII transport, as LMAN1 mutants abolishing carbohydrate binding can still partially rescue secretion. LMAN1/MCFD2-deficient cell lines, overexpression rescue assays, functional secretion assays Blood advances Medium 36490287

Source papers

Stage 0 corpus · 20 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2005 LMAN1 and MCFD2 form a cargo receptor complex and interact with coagulation factor VIII in the early secretory pathway. The Journal of biological chemistry 114 15886209
2005 Combined deficiency of factor V and factor VIII is due to mutations in either LMAN1 or MCFD2. Blood 88 16304051
2006 Cargo selectivity of the ERGIC-53/MCFD2 transport receptor complex. Traffic (Copenhagen, Denmark) 66 17010120
2010 Molecular basis of LMAN1 in coordinating LMAN1-MCFD2 cargo receptor formation and ER-to-Golgi transport of FV/FVIII. Blood 50 20817851
2007 The sugar-binding ability of ERGIC-53 is enhanced by its interaction with MCFD2. Blood 46 18056485
2008 New insights into multiple coagulation factor deficiency from the solution structure of human MCFD2. Journal of molecular biology 31 18590741
2009 EF-hand domains of MCFD2 mediate interactions with both LMAN1 and coagulation factor V or VIII. Blood 23 20007547
2010 Crystal structure of the LMAN1-CRD/MCFD2 transport receptor complex provides insight into combined deficiency of factor V and factor VIII. FEBS letters 22 20138881
2007 Deletion of 3 residues from the C-terminus of MCFD2 affects binding to ERGIC-53 and causes combined factor V and factor VIII deficiency. Blood 20 17971482
2005 Mutations in the MCFD2 gene and a novel mutation in the LMAN1 gene in Indian families with combined deficiency of factor V and VIII. American journal of hematology 20 16044454
2018 Analysis of MCFD2- and LMAN1-deficient mice demonstrates distinct functions in vivo. Blood advances 19 29735583
2022 LMAN1-MCFD2 complex is a cargo receptor for the ER-Golgi transport of α1-antitrypsin. The Biochemical journal 16 35322856
2007 Mutations in the MCFD2 gene are predominant among patients with hereditary combined FV and FVIII deficiency (F5F8D) in India. Haemophilia : the official journal of the World Federation of Hemophilia 14 17610559
2011 Analysis of newly detected mutations in the MCFD2 gene giving rise to combined deficiency of coagulation factors V and VIII. Haemophilia : the official journal of the World Federation of Hemophilia 12 21492322
2023 Separate roles of LMAN1 and MCFD2 in ER-to-Golgi trafficking of FV and FVIII. Blood advances 10 36490287
2022 Effect of co-overexpression of the cargo receptor ERGIC-53/MCFD2 on antibody production and intracellular IgG secretion in recombinant Chinese hamster ovary cells. Journal of bioscience and bioengineering 8 35963666
2020 Crystallographic snapshots of the EF-hand protein MCFD2 complexed with the intracellular lectin ERGIC-53 involved in glycoprotein transport. Acta crystallographica. Section F, Structural biology communications 8 32356523
2008 The first case of combined coagulation factor V and coagulation factor VIII deficiency in Poland due to a novel p.Tyr135Asn missense mutation in the MCFD2 gene. Blood coagulation & fibrinolysis : an international journal in haemostasis and thrombosis 8 18685427
2012 Unveiling the unfolding pathway of F5F8D disorder-associated D81H/V100D mutant of MCFD2 via multiple molecular dynamics simulations. Journal of biomolecular structure & dynamics 5 22208273
2024 RNAi targeting LMAN1-MCFD2 complex promotes anticoagulation in mice. Journal of thrombosis and thrombolysis 1 39222205

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