CXXC to unmethylated CpG DNA: nine mapped contact residues, and an affinity that spans 44 uM to 33 nM depending on the construct, the DNA and the salt
also called MLL CXXC-DNA · 2KKF · C1188D · 44 uM
The fusion protein finds its genes by gripping DNA at unmethylated CpG sites with a small zinc-containing module. How tightly it grips depends enormously on how it is measured - published values run from 44 micromolar to 33 nanomolar - so this entry gives the contact residues and points at the measurement series rather than quoting one number.
The same thing, for a scientist
PDB 2KKF is the solution NMR structure of the KMT2A CXXC domain bound to a palindromic CpG oligonucleotide; the deposited protein entity aligns to Q03164 1147-1203 against the fetched UniProt sequence. NMR titration of that construct gives Kd = 44 uM against a 12-bp single-CpG palindrome in 300 mM NaCl. That figure is the extreme low-affinity point of a nine-measurement series for the same contact spanning to 33 nM - see ana-cxxc-dna-affinity-ladder, added 2026-09-21 - and this entry no longer quotes it as the affinity. Alanine substitution of the electrostatic contacts R1154, K1185 and K1193 costs 9- to 15-fold; Q1187A, which abrogates a specific contact to the guanine base, costs 6-fold; R1150A and L1197A cost 5- and 4-fold; M1200A slightly improves binding, consistent with relief of a steric clash. C1188D, designed to introduce electrostatic repulsion with the DNA backbone, abolishes binding and is the mutation used to test CXXC function in MLL-AF9 leukaemogenesis. The domain lies about 250 residues N-terminal to the breakpoint cluster region and is retained in every leukaemogenic KMT2A fusion.
Contact residues, and how strong this actually is
KMT2A (Q03164): R1150, S1152, R1154, K1185, Q1187, C1188, K1193, L1197, M1200 - all within the crystallised span 1147-1203 1. S1152A could not be expressed solubly, so its contribution is inferred from the structure only.
DNA: a palindromic CpG-containing duplex, CCCTGCGCAGGG, with a single CpG; the specificity determinant is the unmethylated CpG, read in part through Q1187 to the guanine base 1.
CORRECTED 2026-09-21. This section previously read '44 uM is a weak interaction ... three orders of magnitude weaker than the menin contact ... a module that binds its ligand at 44 uM cannot be the sole determinant of targeting.' That comparison is withdrawn [E1 counter].
Ten published measurements of this contact span 44 uM to 33 nM. Sorted by affinity, every adjacent step is 5.5-fold or smaller except the last, which is 19-fold and is the only unresolved one:
| construct | tag | method | DNA | salt | Kd |
|---|---|---|---|---|---|
| 1147-1203 | none | NMR titration | 12 bp, 1 CpG | 300 mM NaCl | 44 uM |
| 1147-1203 | cleaved | ITC | 12-16 bp, 2-3 CpG (Hoxa9) | 300 mM NaCl | 3.5-32 uM across five oligos |
| CXXC, synthesised, no tag | none | EMSA | 12 bp, 1 CpG | 150 mM NaCl | 5.7 uM |
| 1146-1214 | cleaved | ITC | 12 bp, 1 CpG | 250 mM NaCl | 4.3 uM |
| 1147-1203 | GST | fluorescence polarisation | 12 bp, 2 CpG | 150 mM KCl | 0.64 uM |
| 1147-1244 | GST | SPR, DNA immobilised | 52 bp, 6 CpG | 150 mM NaCl | 33 nM |
The 44 uM measurement is the least physiological point of the series on every axis at once: shortest construct, weakest DNA ligand, and twice nuclear ionic strength. The same domain, tagless, against the same class of ligand at 150 mM NaCl binds at 5.7 uM - so halving the salt alone is worth roughly eight-fold. And the menin comparison it was set against - 53 nM for MBM1 - was measured in 50 mM NaCl, one sixth the salt. On the nearest-to-physiological TAGLESS pair (5.7 uM against 53 nM) the ratio is 108-fold, not a thousand-fold, and the menin number still enjoys a third of physiological salt; on Risner's tagged pair it is twelve-fold, and on Birke's geometry the CXXC contact is the tighter of the two. See ana-cxxc-dna-affinity-ladder and ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt.
What survives, on better grounds. The conclusion that this domain does not determine where the fusion goes is unaffected, because it never needed the affinity to be weak. In the MLL-AF9 CXXC swap series the CGBP/CXXC1 domain binds DNA more tightly than the DNMT1 domain and cannot support leukaemogenesis while the weaker DNMT1 domain can, and every swapped domain - including two that bind no DNA detectably - still targeted the fusion to Hoxa9 by ChIP [E1 counter]. The domain reads a two-base motif with no measured readout of flanking bases, so it cannot specify an address at any affinity. See mechanism-the-cxxc-address-argument-is-about-specificity-not-affinity.
Relation to the existing CXXC entries
The graph holds target-kmt2a-cxxc-dna-interface (a druggability dossier), localization-cxxc-cpg-address and mechanism-fusion-dna-targeting-cxxc-athooks. This entry supplies the contact residues and the structural provenance; ana-cxxc-dna-affinity-ladder owns the affinity series and interface-kmt2a-mt-bcb-domain-1147-1244 owns the longer MT/BCB construct.
CORRECTED 2026-09-21. This section previously read: the 44 uM number adds a constructive angle - a competitor does not need to be potent in absolute terms to displace a 44 uM binder, it needs to be selective. That is withdrawn. If the species in the cell is nearer the multivalent 33 nM end of the series, a DNA-competitive ligand must be nanomolar to compete. The potency bar rises by up to three orders of magnitude and the low-hanging-fruit framing goes with it. The dossier's verdict of a flat charged protein-DNA interface with zero chemical matter is unaffected and if anything is reinforced.
| partner a | gene-kmt2a |
|---|---|
| partner b | unmethylated CpG DNA |
| motif a | KMT2A ZF-CXXC domain; crystallised span 1147-1203; UniProt CXXC-type zinc-finger feature 1147-1195 (live fetch 2026-09-21); the MT/BCB domain of the functional literature is 1147-1244 |
| motif b | palindromic duplex containing a single unmethylated CpG |
| fate in fusion | retained - the domain lies about 250 residues N-terminal to the breakpoint cluster region and is present in every leukaemogenic KMT2A fusion |
| affinity context | WITHDRAWN AND REPLACED 2026-09-21. This field formerly read: '44 uM is roughly 1,000-fold weaker than the menin-MBM1 contact (53 nM) and 250,000-fold weaker than the AF9 AHD-AF4 contact (0.17 nM) made by the same complex.' The CXXC figure was measured at 300 mM NaCl and the menin figure at 50 mM NaCl, on interactions with opposite salt sensitivities. On the nearest-to-physiological tagless pair (5.7 uM at 150 mM NaCl against 53 nM at 50 mM NaCl) the ratio is 108-fold, and on the tagged pair (0.64 uM at 150 mM KCl) it is 12-fold; on the MT-domain SPR geometry at 150 mM NaCl the CXXC contact is tighter than the menin contact. No published pair of measurements supports a stated fold-difference between these two contacts. See ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt. |
| druggability | See target-kmt2a-cxxc-dna-interface for the full dossier. CORRECTED 2026-09-21: this field formerly said a competitor needs selectivity rather than raw potency because the natural ligand binds at 44 uM. If the operative species is nearer the multivalent 33 nM end of the affinity series, a DNA-competitive ligand needs nanomolar potency AND family selectivity. The potency bar rises by up to three orders of magnitude. |
| resolution note | PDB metadata and entity sequences fetched from data.rcsb.org 2026-09-20; the CXXC span aligned to UniProt Q03164 fetched the same day |
| paralogue selectivity note | Xu et al. measured and compared the DNA-binding selectivities of all known human CXXC domains and classified them by selectivity rather than by function, so CXXC domains are not interchangeable DNA readers. |
| kd NM note | DELIBERATELY NULL as of 2026-09-21. A single number for this contact is not defensible: ten published measurements span 44 uM to 33 nM across construct, tag, DNA ligand, ionic strength and readout. The nearest-to-physiological tagless value is 5.7 uM at 150 mM NaCl (PMID 34280370). The full series with conditions is in ana-cxxc-dna-affinity-ladder. The former value in this field was 44000. |
| uniprot zf cxxc feature | 1147-1195, fetched live from rest.uniprot.org 2026-09-21; the 1242 figure formerly recorded here was wrong |
| affinity series owner | ana-cxxc-dna-affinity-ladder |
interface residues a
[ "KMT2A R1150", "KMT2A S1152", "KMT2A R1154", "KMT2A K1185", "KMT2A Q1187", "KMT2A C1188", "KMT2A K1193", "KMT2A L1197", "KMT2A M1200" ]
hot spot residues
[ "KMT2A K1185", "KMT2A K1193", "KMT2A R1154" ]
kd measurements
[
{
"ligand": "wild-type CXXC 1147-1203 + palindromic 12-mer CCCTGCGCAGGG, 1 CpG",
"kd_nM": 44000,
"method": "NMR titration",
"salt_mM": 300,
"salt_species": "NaCl",
"tag": "none",
"source_pmid": "20010842"
},
{
"ligand": "CXXC 1147-1203 + five Hoxa9 12-16mers",
"kd_nM": "3500-32000",
"method": "ITC",
"salt_mM": 300,
"salt_species": "NaCl",
"tag": "GST cleaved",
"source_pmid": "18483194"
},
{
"ligand": "CXXC 1146-1214 + 12-mer, 1 CpG",
"kd_nM": 4300,
"method": "ITC",
"salt_mM": 250,
"salt_species": "NaCl",
"tag": "lipoyl, TEV-cleaved",
"source_pmid": "16990798"
},
{
"ligand": "chemically synthesised untagged CXXC + 12-mer palindrome GCCACCGGTGGC, 1 CpG",
"kd_nM": 5700,
"kd_se_nM": 200,
"method": "EMSA, Hill fit, N = 7",
"salt_mM": 150,
"salt_species": "NaCl",
"tag": "none - chemically synthesised",
"source_pmid": "34280370"
},
{
"ligand": "GST-CXXC 1147-1203 + 12-mer GGGTCGCGGGAG, 2 CpG",
"kd_nM": 640,
"method": "fluorescence polarisation",
"salt_mM": 150,
"salt_species": "KCl",
"tag": "GST uncleaved",
"source_pmid": "23990460"
},
{
"ligand": "GST-MT 1147-1244 + 52-bp CpG52, 6 CpG, DNA immobilised",
"kd_nM": 33,
"method": "SPR, k_d/k_a from initial rates",
"salt_mM": 150,
"salt_species": "NaCl + 6 mM MgCl2",
"tag": "GST uncleaved",
"source_pmid": "11842107"
},
{
"ligand": "R1154A / K1185A / K1193A",
"fold_loss": "9 to 15",
"method": "NMR titration",
"source_pmid": "20010842"
},
{
"ligand": "Q1187A",
"fold_loss": "6",
"method": "NMR titration",
"source_pmid": "20010842"
},
{
"ligand": "R1150A",
"fold_loss": "5",
"method": "NMR titration",
"source_pmid": "20010842"
},
{
"ligand": "L1197A",
"fold_loss": "4",
"method": "NMR titration",
"source_pmid": "20010842"
},
{
"ligand": "C1188D",
"fold_loss": "binding abolished",
"method": "NMR titration",
"source_pmid": "20010842"
},
{
"ligand": "M1200A",
"fold_loss": "moderate GAIN of affinity",
"method": "NMR titration",
"source_pmid": "20010842"
}
]structures
[ "2KKF", "4NW3", "2J2S", "2JYI" ]
structure details
[
{
"pdb": "2KKF",
"title_fetched": "Solution structure of MLL CXXC domain in complex with palindromic CPG DNA",
"method": "NMR",
"resolution_A": null,
"contents": "Q03164 1147-1203 + 12-mer DNA duplex",
"fetched": "2026-09-20"
},
{
"pdb": "4NW3",
"title_fetched": "Crystal structure of MLL CXXC domain in complex with a CpG DNA",
"method": "X-ray",
"resolution_A": 2.82,
"contents": "Q03164 CXXC (His-TEV tagged construct) + 12-mer DNA duplex GCCATCGATGGC",
"fetched": "2026-09-20"
},
{
"pdb": "2J2S",
"title_fetched": "Solution structure of the nonmethyl-CpG-binding CXXC domain of the leukaemia-associated MLL histone methyltransferase",
"method": "NMR",
"resolution_A": null,
"contents": "free CXXC domain, 72-mer entity",
"fetched": "2026-09-20"
},
{
"pdb": "2JYI",
"title_fetched": "Solution structure of MLL CXXC domain",
"method": "NMR",
"resolution_A": null,
"contents": "free CXXC domain, 57-mer entity; RCSB records no primary citation PMID or DOI for this entry",
"fetched": "2026-09-20"
}
]menin mbm1 kd conditions
{
"_why": "Added by king sweep 2026-09-21 (Q82 corrections-owed #3). This entry quotes the menin-MBM1 53 nM figure. That number is NOT comparable to the KMT2A CXXC-DNA affinities without stating its buffer, and the omission is how a 1,300-fold 'discrepancy' was manufactured.",
"value": "53 nM",
"motif": "KMT2A MBM1, residues 4-15, FITC-labelled peptide at 15 nM",
"method": "fluorescence polarisation",
"buffer": "50 mM Tris, pH 7.5, 50 mM NaCl, 1 mM DTT",
"nacl_mM": 50,
"companion_itc": "72 nM by ITC, 50 mM phosphate pH 7.5, 50 mM NaCl, 1 mM beta-mercaptoethanol, 27 C",
"source_pmid": "20961854",
"source_quote": "\"in the FP buffer (50 mM Tris, pH 7.5, 50 mM NaCl, 1 mM DTT) used for binding experiments (Kd measurements)\" -- Grembecka 2010 JBC, Methods, fetched and read from PMC3003368 on 2026-09-21",
"do_not_compare_naively": {
"against": "KMT2A CXXC-DNA Kd 44 uM (PMID 20010842, Cierpicki 2010)",
"because": "that measurement was made at 300 mM NaCl -- verbatim from its Methods, \"50 mM phosphate buffer (pH 7.1), 300 mM NaCl, 2 mM DTT at 25 C\" (PMC2908503, read 2026-09-21) -- SIX-FOLD more salt, on a salt-sensitive protein-DNA contact, against a hydrophobic peptide pocket that is not.",
"best_matched_pair": "44 uM at 300 mM NaCl vs the 5.7 uM tagless CXXC measurement at 150 mM NaCl gives 108-fold against menin, not ~1,000-fold",
"see": "ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt"
},
"added": "2026-09-21"
}cxxc dna kd conditions
{
"_why": "Added by king sweep 2026-09-21 (cycle 46). This entry quotes the KMT2A CXXC-DNA 44 uM. That number is a property of five choices, not of the molecule, and quoting it bare is how this graph manufactured a 1,300-fold 'discrepancy' with the menin contact.",
"value": "44 uM",
"construct": "isolated CXXC domain, KMT2A 1147-1203, untagged, 15N-labelled",
"ligand": "DNA pal oligonucleotide, one CpG element",
"method": "NMR titration",
"buffer": "50 mM phosphate, pH 7.1, 300 mM NaCl, 2 mM DTT, 25 C",
"nacl_mM": 300,
"source_pmid": "20010842",
"source_quote": "\"Determination of the dissociation constants by NMR for the CXXC domain (MLL 1147-1203) and mutants was performed by titration of 15N-labeled protein with concentrated solutions of DNA pal oligonucleotide. Experiments were carried out in 50 mM phosphate buffer (pH 7.1), 300 mM NaCl, 2 mM DTT at 25 C.\" -- Cierpicki 2010 NSMB, Methods, fetched and read from PMC2908503 on 2026-09-21. The result sentence reads \"The wild type protein yielded a Kd of 44 uM\".",
"this_is_the_high_salt_end": "300 mM NaCl is roughly twice physiological ionic strength and SIX TIMES the 50 mM the menin-MBM1 53 nM was measured in. The same domain measured tagless at 150 mM NaCl gives 5.7 uM -- 7.7-fold tighter from salt alone.",
"do_not_compare_naively": {
"against": "menin-MBM1 53 nM (PMID 20961854), measured at 50 mM NaCl by FP",
"best_matched_pair": "108-fold, not ~1,000-fold",
"see": "ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt"
},
"added": "2026-09-21"
}- Functional substitution: the DNMT1 CXXC domain can replace the MLL CXXC in MLL-AF9 leukaemogenesis (Risner 2013, PMID 23990460), so blocking this specific surface invites substitution rather than loss of function.
- Paralogue breadth: CXXC domains are shared by KMT2B, DNMT1, TET1, TET3, CXXC1, KDM2A, KDM2B, MBD1, CXXC4, CXXC5 and FBXL19 - a family closed at twelve including KMT2A - several of which are essential.
- CORRECTED 2026-09-21. This item formerly read that a 44 uM protein-DNA contact is the least tractable geometry for a small molecule 'even though the low affinity lowers the potency bar'. The second clause is withdrawn: the affinity series runs to 33 nM and the potency bar may be nanomolar. The geometry argument stands and the consolation does not.
- Redundant anchoring: the fusion also carries AT-hooks and is tethered by menin and LEDGF, so removing one anchor may not move occupancy at all - and Risner's swap series shows the fusion still reaches Hoxa9 even with a CpG-binding-dead module.
- Fractional occupancy cannot be computed from a Kd alone - it needs the free nuclear concentration of the fusion, which has never been measured. This entry formerly asked what fraction of CpG-island occupancy is CXXC-driven given a 44 uM affinity; that question is unanswerable as posed and is re-stated in ana-cxxc-occupancy-is-underdetermined-by-every-published-kd.
- What is the affinity of the 1147-1244 MT/BCB construct with no tag at 150 mM salt? It is the only unresolved step in the affinity series and one titration closes it.
- Does C1188D, the mutation that abolishes DNA binding, change fusion residence time on chromatin measurably, or only steady-state occupancy?
- 2JYI has no associated publication in the RCSB record - what is its provenance, and does it agree with 2J2S and 2KKF?
- E1established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.paper
The KMT2A CXXC domain binds the palindromic 12-mer CCCTGCGCAGGG with Kd 44 uM by NMR titration in 50 mM phosphate pH 7.1 with 300 mM NaCl at 25 C; R1154A, K1185A and K1193A cost 9- to 15-fold, Q1187A 6-fold, R1150A 5-fold, L1197A 4-fold, and C1188D abolishes binding.
Structure of the MLL CXXC domain-DNA complex and its functional role in MLL-AF9 leukemia. · Cierpicki T, Risner LE, Grembecka J, et al. · Nat Struct Mol Biol 2010What this evidence cannot show
NMR titration affinities on a short duplex with a single CpG, at 300 mM NaCl - roughly twice nuclear monovalent ionic strength - on a 57-residue construct; a genomic site is a different ligand and a cell is a different buffer. S1152A could not be expressed solubly so its structural assignment is untested. All mutants were confirmed folded by HSQC. THIS 44 uM IS THE EXTREME LOW-AFFINITY POINT OF A NINE-MEASUREMENT SERIES SPANNING TO 33 nM (see ana-cxxc-dna-affinity-ladder, added 2026-09-21) AND MUST NOT BE QUOTED AS THE AFFINITY OF THIS CONTACT. Full text re-read 2026-09-21 via the PMC HTML route (PMC2908503) after Europe PMC fullTextXML returned HTTP 500; the buffer and oligonucleotide sequence are taken from the paper's own Methods. Supplementary Table 1, which holds the paper's ITC values for other oligonucleotides, is behind the Europe PMC open-access gate and was not read.
Evidence that argues against this (1)
- CORRECTED 2026-09-21. This note previously attributed the DNMT1-CXXC substitution result to Allen 2006 (PMID 16990798). Allen 2006 is a solution-structure paper and contains no such experiment - verified by reading its full text this cycle, where the strings 'substitut', 'leukaemogenesis' and 'immortal' occur only in a sequence-conservation sentence and in the reference list. The experiment is Risner 2013 (PMID 23990460), now cited here. Its content: the DNMT1 CXXC domain substitutes functionally for the MLL CXXC in MLL-AF9 leukaemogenesis, so the contact chemistry mapped here is not what the oncoprotein requires - it requires the function of protecting particular CpGs, which another CXXC can supply. And the same paper shows the CGBP/CXXC1 domain binds DNA MORE tightly than DNMT1's and fails, while every swapped domain including two non-binders still targeted the fusion to Hoxa9 by ChIP, so DNA-binding affinity predicts neither function nor localisation here. Risner LE et al. 2013
- E2established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.paper
The MLL CXXC domain is a non-methyl-CpG-binding module, defined structurally in solution independently of the DNA complex.
Solution structure of the nonmethyl-CpG-binding CXXC domain of the leukaemia-associated MLL histone methyltransferase. · Allen MD, Grummitt CG, Hilcenko C, et al. · EMBO J 2006What this evidence cannot show
A free-domain NMR structure; it establishes the fold and the methylation-sensitivity but not the contact register with DNA.
Evidence that argues against this (1)
- The DNA-bound structure identified contacts in the minor groove - R1150, S1152 and L1197 - that the free-domain analysis did not, so a free structure under-reports the interface. Cierpicki T et al. 2010
- E3established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.database
PDB 2KKF contains a KMT2A entity mapping to Q03164 1147-1203 together with a 12-mer palindromic CpG DNA duplex.
PDB 2KKF: Solution structure of MLL CXXC domain in complex with palindromic CPG DNA · Cierpicki T, Risner LE, Grembecka J, et al. · RCSB Protein Data Bank 2010What this evidence cannot show
Title, method and both entity sequences fetched from data.rcsb.org 2026-09-20; the protein span is that cycle's alignment to the UniProt Q03164 sequence, after stripping the GS expression tag. NMR, so no resolution applies. CORRECTED 2026-09-21: this caveat previously said 'the UniProt ZF-CXXC feature extends beyond the crystallised span, to roughly 1242.' Live UniProt Q03164, fetched 2026-09-21 by REST, gives the CXXC-type zinc-finger feature as 1147-1195 - SHORTER than the crystallised span, not longer - with a disordered region annotated at 1200-1375. The 1242/1244 figure belongs to the MT/BCB domain of the 2002 functional literature, not to the UniProt feature; see interface-kmt2a-mt-bcb-domain-1147-1244.
Evidence that argues against this (1)
- Two further NMR structures of the free MLL CXXC domain exist (2J2S and 2JYI) from different groups, and 2JYI has no associated publication, so the deposited structural record for this domain is uneven in provenance. Allen MD et al. 2006
- E4established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.paper
A crystal structure of the MLL CXXC domain bound to CpG DNA exists at 2.82 A (PDB 4NW3), from a systematic comparison of the DNA-binding selectivities of all known human CXXC domains.
DNA Sequence Recognition of Human CXXC Domains and Their Structural Determinants. · Xu C, Liu K, Lei M, et al. · Structure 2018What this evidence cannot show
Entry metadata and both entity sequences fetched from data.rcsb.org 2026-09-20; the paper has no PMCID and its full text was not retrievable this cycle, so the selectivity classification is taken from the abstract fetched from PubMed. At 2.82 A the crystal is lower resolution than the peptide co-crystals elsewhere in this beat.
Evidence that argues against this (1)
- The paper's own framing is that human CXXC domains differ in DNA selectivity, which cuts against treating any one CXXC domain as interchangeable with another - and therefore against the simple reading of the DNMT1-CXXC substitution result. Xu C et al. 2018
- E5moderate confidence 3 of 5: One strong study, or several smaller consistent ones.paper
A tagless, chemically synthesised MLL1 CXXC domain binds a 12-bp single-CpG palindrome with Kd 5.7 +/- 0.2 uM by EMSA at 150 mM NaCl, and the same paper reports dimethyl fumarate inhibiting the CXXC-DNA complex with a relative IC50 of 0.91 +/- 0.09 uM in a single-molecule assay while the metabolite monomethyl fumarate is inactive at 10 uM.
Dynamics and inhibition of MLL1 CXXC domain on DNA revealed by single-molecule quantification. · Liang L, Ma K, Wang Z, et al. · Biophys J 2021What this evidence cannot show
EMSA rather than a solution-equilibrium method, run on ice, so the roughly eight-fold difference from the 44 uM NMR figure confounds ionic strength with method and temperature; the construct span is in an unretrieved supplementary table. The DMF figure is a RELATIVE IC50 from a four-parameter logistic fit to single-molecule unfolding times, with no counter-screen against any other CXXC domain and no cellular experiment. Full text read 2026-09-21 via the PMC HTML route (PMC8392069). This paper was invisible to this project's PubMed sweeps because MLL[tiab] does not match the token MLL1.
Evidence that argues against this (1)
- DMF covalently modifies more than 2,400 cysteines and the CXXC fold coordinates zinc through eight of them, so sub-micromolar inhibition is the expected generic result for a cysteine-reactive electrophile rather than evidence that this interface is ligandable. Every other member of the twelve-protein CXXC family would be expected to behave the same way, and none was tested. Risner LE et al. 2013
CXXC 1147-1203 on unmethylated CpG DNA, Kd 44 uM
supplies the affinity and nine contact residues that dossier lacked
the residue-level basis of the CpG address
- supports →Step 1 — How the fusion finds its genes: AT-hooks and the CXXC domain reading unmethylated CpG
quantifies how weak the CXXC contribution is on its own
retained in the index fusion
- ← related toThe human CXXC/CpG-reading family is exactly twelve proteins, and TET2 is not one of them
The KMT2A CXXC at Q03164 1147-1195 is one of twelve human instances of the same fold; the other eleven are its selectivity problem.
One of the eleven other human CXXC domains any ligand for this interface would have to discriminate against.
Head-to-head: MLL1-CXXC tolerates no cytosine modification, TET3-CXXC tolerates 5mC, 5fC and 5caC and does not require CpG context - the sharpest published contrast between the index domain and a family member.
The family member whose reader chemistry is closest to the index domain - non-methylated CpG, blocked by methylation - and therefore the hardest selectivity problem for a CXXC-directed ligand.
Its CXXC domain was swapped into MLL-AF9 in place of this interface's domain and abrogated leukaemogenesis - but did not bind DNA, so the arm is a control rather than a test.
One of the eleven other human CXXC domains a ligand for this interface must discriminate against, and the one that is almost nothing but the domain.
One of the eleven other human CXXC domains a ligand for this interface must discriminate against.
The one heterologous CXXC domain that can replace this one in MLL-AF9 and still run a leukaemia - the tightest functional constraint on what this interface has to do.
One of the eleven other human CXXC domains a ligand for this interface must discriminate against; tandem CXXC-PHD architecture like KDM2A and KDM2B.
The eleven other domains that share this fold have never been measured alongside it in a cell.
- ← related toIs the CpG address a contested resource? On current evidence, no - and the beat that went looking says so
If the address is not contested, this weak (~44 uM) interface is buttressed by the AT-hooks and the menin-LEDGF tether rather than by affinity.
Tests whether this weak (~44 uM) interface is displaceable in a cell by a rival reader.
Records the unreconciled affinity gap: 33 nM by SPR on a GST-MT fusion versus ~44 uM by NMR on an isolated domain.
If the shared 'DNA-binding domain' is the CXXC, decitabine is an accidental low-potency CXXC-directed agent; C1188D is the falsifier.
Withdraws the entry's headline comparison: 44 uM is the extreme low-affinity point of a nine-measurement series spanning to 33 nM, measured at 300 mM NaCl against the shortest construct and a single-CpG 12-mer.
The structural entry describes 1147-1203; this entry describes the 98-residue construct the functional literature has always used, and corrects the UniProt ZF-CXXC boundary from 'roughly 1242' to the live 1147-1195.
Supplies the sequence dependence the structural entry's single-oligonucleotide affinity cannot show.
The entry's 'three orders of magnitude weaker than the menin contact' compares a 300 mM NaCl measurement with a 50 mM NaCl one.
The entry's open question asking what fraction of occupancy is CXXC-driven given a 44 uM affinity is unanswerable: both the Kd and the free nuclear concentration are unmeasured.
Withdraws the entry's claim that a 44 uM module cannot be the sole determinant of targeting because it is weak, replacing it with a specificity argument that does not depend on the magnitude.
The same paper supplies 4NW3, which that entry already records from RCSB metadata.
The only published chemical inhibition of the interface this entry maps, from the same paper that supplies the near-physiological 5.7 uM affinity.