four designed enzyme chemistries can plausibly engage the widest range of HNE-derived Michael adducts and crosslinks, using three independent methods so the answer doesn’t rest on one modeling assumption. All three measure geometric/pose plausibility of reaching the catalytic anchor — not catalytic turnover.
The ranking
AKR1C1 > SPRTN > AAG ≫ TDP2 (geometric ≤5 Å gate):
Enzyme Coverage Passing / target-bond-present Median reactive→anchor Native breadth AKR1C1 0.76 19 / 25 3.57 Å broad promiscuous reductase SPRTN 0.67 12 / 18 3.23 Å broad, sequence-independent AAG 0.50 6 / 12 5.00 Å broad among base lesions TDP2 — (0/12) 0 target-bond-present n/a narrow specialist
This computational ordering is concordant with the enzymes’ documented native substrate breadth (three broad enzymes vs. one narrow specialist).
The two findings that matter most for honesty
AKR1C1 is “broadest” only by docking proximity. Under the mechanistically-strict reduction gate (carbonyl within hydride distance of NADP-C4N and oriented to the Tyr55 proton donor), its coverage collapses from 0.76 to 0.20 (5/25). The limiting factor is Tyr55 geometry, not the hydride distance — many adducts reach the nicotinamide but don’t simultaneously present the carbonyl to the acid. So its docking breadth is wide while its productive-reduction breadth is narrow — and, per the earlier boundary probe, it offers no crosslink-release chemistry at all.
TDP2’s 0/12 is a genuine mechanistic negative, not a design failure. No HNE or base lesion carries a 5′-phosphotyrosyl bond, so none presents TDP2’s target bond — exactly matching its specialized native role.
Read mechanistically, SPRTN is the enzyme best positioned to engage the widest range of bona fide crosslinks (as opposed to free/residual carbonyls): its sequence-independent protease chemistry reaches the scissile bond for both small electrophiles (in the top-scoring pose) and bulky HNE-DPCs (in a sub-optimal pose within ~0.7 kcal/mol).