Krey 2016 J Cell Biol (PMC5119939) — not retrieved as paper note
FFT-EM
Espin-crosslinked spacing
~12 nm
Review consensus (Krey 2016 cites)
for comparison
Red flags in current h09 Phase 4d model
Model constant
Value
Problem
WH2_KD_GACTIN_M = 100 nM
literature Kd
Partially defensible for WAVE-like WH2 (50–100 nM). For WASP-type (which is what WH2-family SAPs typically use) real value is 200 nM – 3 μM. Model comment should read “50–500 nM depending on construct”.
WH2_KD_FACTIN_M = 5 μM
”estimated 5–25× weaker”
LOAD-BEARING RISK. Zero primary measurements exist. Closest analog (Tβ4 × F-actin) = 5–10 mM = 1,000–2,000× weaker than model’s 5 μM. Structurally disfavored (binding site buried in F-filament). Avidity from multi-WH2 on RADA16 scaffold could compensate in principle; Phase 2c wet-lab bundling assay is the only resolution.
STEREOCILIA_INTER_FILAMENT_NM = 12.0
espin-specific
Native OHC uses plastin-1/fimbrin → real spacing 7.9–9.7 nm (Krey 2016). For STRC exoprosthesis modeling, use 9 nm as default.
Critical takeaway
Central unvalidated assumption of h09: WH2 can side-bind F-actin at a therapeutically relevant Kd. Only possible via avidity (multiple WH2 copies per RADA16 fibril contacting the same F-actin surface). Single-molecule Kd at 5 μM is speculative optimism. Phase 2c wet-lab actin-bundling assay (co-sedimentation + TIRF colocalisation) becomes the primary risk gate, not AF3 triple-complex.