RADA16 / SAP geometry + in vivo kinetics — validated parameters
Source agent: Domain 4 (Sonnet 4.6), 2026-04-23. Consumer: hydrogel_phase4d_factin_bundling_model.py (scaffold geometry) + hydrogel_phase4e_cochlear_pkpd.py (gel clearance).
Parameter table
| Parameter | Literature value | Source | Status | Used in |
|---|---|---|---|---|
| Antiparallel β-sheet H-bond spacing | 4.7 Å (0.47 nm) | Cormier 2012 Biomacromolecules (WAXD, paywalled abstract); amyloid standard | ✅ primary | Phase 4d fibril math |
| Peptides per nm fibril axis (derived) | 4.26/nm (bilayer: 2 peptides per 0.47 nm step) | 2005-yokoi-kinoshita-zhang-rada16-reassembly + Paravastu 2014 double-sheet model | ✅ derived | Phase 4d RADA16_MONOMERS_PER_NM_FIBRIL |
| Single nanofiber dimensions (tape) | 3 nm wide × 2.5 nm high (flat, 2 β-sheets, alanine core) | Paravastu 2014 ACS Nano (PMC3946435, solid-state NMR) | ✅ primary | cross-section context |
| Bundle diameter (EM observations) | 10–20 nm (aggregated ~3–7 individual nanofibers) | 1993-zhang-eak16-rada16-spontaneous-assembly Zhang 1993 PNAS 90:3334 | ✅ primary | bundle vs fibril distinction |
| Mean fibril length (AFM intact fibers) | 615 ± 104 nm | 2005-yokoi-kinoshita-zhang-rada16-reassembly | ✅ primary | — |
| Mean fibril length (pH 4.5, end-to-end assembly) | 296 ± 131 nm | PMC3318125 | ✅ primary | pH-dependent |
| Observed fibril length range | 200 nm to few μm | 2005-yokoi-kinoshita-zhang-rada16-reassembly | ✅ primary | Phase 4d fibril_length_nm = 1000 upper-range |
| β-strand axial tilt | ~35° from fibril axis (2-residue registry shift, Class 3R2) | Paravastu 2014 | ✅ primary | — |
In vivo PK (assembled RADA16 gel)
| Condition | t½ | Source | Notes |
|---|---|---|---|
| Rat liver injury (uncrosslinked RADA16) | 3–7 days visible; dissolved by 14 days | 2021-frontiers-rada16-clinical-review (PMC8216384) | Direct histology |
| Rat TBI brain (uncrosslinked) | Visible at 7 days; absent at 21 days | 2021-frontiers-rada16-clinical-review | Direct histology |
| Crosslinked RADA16 variants | 61% degraded at 35 days | 2021-frontiers-rada16-clinical-review | Extended persistence |
| Derived assembled gel k_dissolution | 0.002–0.004/h (t½ 7–14 days) | 0.693 / (7–14 days × 24 h) | calculation |
| Free monomeric 16-mer in serum | ~30 min plausible (not measured directly for RADA16) | General small-peptide serum stability | inferred |
Tail-length modification limits (h09-critical)
Empirical assembly limit for appended sequences onto RADA16:
| Appended length | Assembly behavior | Source |
|---|---|---|
| ≤12 residues | β-sheet assembly preserved | 2021-frontiers-rada16-clinical-review (Frontiers 2021 review, PMC9739689 citations) |
| 12–50 residues | Monotonic reduction in viscoelasticity / β-sheet signal | Same review |
| ALK-tagged RADA16 (+3 hydrophobic residues) | Rapid collapse to spherical aggregates | Same review |
| >50 aa globular domain | Never tested in published literature | — |
| Our tail91 construct: 118 aa | UNCHARACTERIZED, ~10× beyond empirical limit | — |
Red flags in current h09 Phase 4d/4e model
| Model constant | Value | Problem |
|---|---|---|
RADA16_MONOMERS_PER_NM_FIBRIL = 4.35 | comment “Zhang MIT” | Value defensible (2% off 4.26). Comment misleading — Zhang 1993 provides no such number. Correct citation: Cormier 2012 (WAXD H-bond spacing) + Paravastu 2014 (bilayer model). |
fibril_length_nm = 1000 | inline | Defensible as upper-range AFM value; mean ~615 nm (Yokoi 2005). Flag as upper bound. |
RADA16_FIBRILS_PER_CROSSSECTION = 5 (avg) | — | Geometrically plausible (10–20 nm bundle / 3 nm tape width = 3–7 fibers/bundle). Not directly stated in any paper. |
K_PROTEOLYSIS = 1.4/h (t½ 30 min) | “conservative” | Wrong regime. For assembled gel, in vivo t½ is 7–14 days → k ≈ 0.002–0.004/h. Model is 350–700× faster than reality in the error-safe direction (overestimates clearance). Need to split: k_gel_dissolution (slow, scaffold at delivery site) vs k_free_monomer_proteolysis (fast, for peptide monomers that haven’t assembled). |
| 118 aa tail on RADA16 | core design assumption | NOT IN CHARACTERIZED PARAMETER SPACE. Empirical limit for sequence modification preserving assembly is ~12 residues. Our tail is 10× beyond. Rigorous path: 9:1 molar blend (plain RADA16 : RADA16-tail91) where plain scaffold carries assembly and tail91 displays WH2/TMEM145-binder on surface. Documented in literature for similar hybrids (Gelain osteogenic). Must be added to Phase 4e design. |
Critical implication
h09 Phase 3b PEPTIDE_TAIL91 cannot be assumed to self-assemble at 100% composition. Every dose calculation in Phase 4e that treats the entire administered peptide as contributing to the functional scaffold is 10× too optimistic if a 9:1 blend is used (only 10% of mass is functional). Alternatively, if pure tail91 is used, assembly likely fails entirely.
Peptide-design / sequence-search references (added 2026-04-25)
For h09 sequence-design loops (e.g., redesigning the WH2-display tail or testing residue-conservation series on the RADA core), the canonical de novo / nature-inspired-algorithm reference is now ingested:
| Reference | Role | Local note |
|---|---|---|
| Schneider (Ed.) 2014, De novo Molecular Design, Wiley-VCH | de novo design, FBDD, peptide design by SME/PSO/ACO, peptide stability mods | 2014-schneider-de-novo-molecular-design-book |
Recipes available:
- Recipe — Ant Colony Optimization for Peptide Sequence Design — verbatim §18.3 pseudocode; 89%/95% accuracy on the MHC-I octapeptide validation case.
- Recipe — Therapeutic Peptide Stability Modifications — cyclization, stapling, end-capping, glycosylation, PASylation (§18.4).
Reference data (P0):
- Amino Acid Physicochemical Distance Matrix Grantham Modified — verbatim Table 18.1 (modified Grantham, row-scaled to [0,1]). Used to parameterize SME mutation step σ for h09 ablation series.
Connections
[part-of]_hub (literature-params)[see-also]cochlear-pkpd (gel dissolution constant conflicts with K_PROTEOLYSIS)[see-also]actin-kinetics (avidity argument rests on multi-WH2 per fibril)[applies]STRC Synthetic Peptide Hydrogel HTC[applies]index[see-also]2014-schneider-de-novo-molecular-design-book[see-also]Recipe — Ant Colony Optimization for Peptide Sequence Design[see-also]Recipe — Therapeutic Peptide Stability Modifications[see-also]Amino Acid Physicochemical Distance Matrix Grantham Modified