Aldolase A - 3D Mechanism Simulator

Loading structural data… (1ZAI · 6ALD · 1ADO)

References & Databases

Structure — covalent imine and Michaelis states 1ZAI / 1ZAH = rabbit muscle class-I aldolase trapping the Schiff-base (imine) intermediate (St-Jean M, Lafrance-Vanasse J, Liotard B, Sygusch J, J Biol Chem 280:27262–27270, 2005, PDB 1ZAI · PMID:15870069 · DOI:10.1074/jbc.M502413200). 6ALD = the cleavage-defective K146A mutant trapping its natural substrate as a Michaelis (non-covalent) complex (Choi KH, Mazurkie AS, Morris AJ, Utheza D, Tolan DR, Allen KN, Biochemistry 38:12655–12664, 1999, PDB 6ALD · PMID:10504235). 1ADO = native rabbit enzyme (Blom N & Sygusch J, Nat Struct Biol 1997, PDB 1ADO · PMID:8989320). Mechanism record: M-CSA Entry 222 (class-I aldolase, UniProt P00883).
① F1,6BP Michaelis complex — re-anchored [schematic re-anchor = frame offset] The 6ALD substrate co-crystallizes with its C2 carbonyl ≈ 7.27 Å from Lys229-NZ (6ALD frame). Re-measuring that same pose in the 1ZAI imine frame — where the lysine seat differs — reads ≈ 6.83 Å (drawn as ≈ 6.8 Å): a frame difference, not a rotation; no carbonyl-environment superposition was applied. Ser271-OG…P1 = 3.8 Å seats the C1-phosphate; the deposited C6-phosphate sits with Arg303/Arg42/Lys41.Source: measured — 6ALD substrate pose C2…Lys229-NZ = 7.27 Å in the 6ALD frame (PDB 6ALD); 6.83 Å (drawn ≈ 6.8 Å) = the same pose re-read in the 1ZAI imine frame — own common-frame value, frame offset [schematic], not a rotation (Why-is-the-Michaelis card) · Ser271-OG…P1 = 3.8 Å, common-frame measured
② Schiff base (imine) Lys229 ε-amine attacks the C2 carbonyl to give the protonated imine — measured C2–NZ = 1.30 Å in 1ZAI; a contact network (Lys146, Glu187, Asp33) tethers the C4-hydroxy (2.59 / 3.05 / 4.64 Å). Condensation water leaves as a byproduct [schematic].Source: measured in PDB 1ZAI (St-Jean et al. 2005): imine C2–NZ = 1.30 Å · Lys146 / Glu187 / Asp33 to C4-hydroxy = 2.59 / 3.05 / 4.64 Å · leaving condensation water = [schematic] (no crystal)
③ Retro-aldol C3–C4 cleavage (PROPOSAL) [schematic · hypothetical, lit.-based] M-CSA Entry 222 assigns Asp33 (Asp34 in UniProt numbering) as the general base abstracting the C3 proton as the C3–C4 bond breaks; D-G3P (C4–C6) leaves first, the enamine/imine form staying on the DHAP fragment. (M-CSA's role table — reference PDB: DHAP–enamine 2QUT — additionally lists Tyr363 (C3 pro-S removal), Ser300 (enamine stabilizer) and Glu189 (water activation); the C3-proton carrier is contested — St-Jean 2005 routes the C3–C4 cleavage through Glu187 with Lys146 stabilizing the C4-OH.)Source: PROPOSAL — no retro-aldol TS observed (Prop card); Asp33 general base (UniProt Asp34) + G3P-leaves-first order per M-CSA Entry 222 · Glu187-route alternative: St-Jean et al. 2005 (Base-assignment card: complementary, not competing)
④ Imine hydrolysis [schematic] A water (O red · H white) is activated by Glu187 (M-CSA: deprotonates the hydrolytic water attacking the imine carbon); Lys229-NH₂ is regenerated and DHAP dissociates toward the TIM (triose phosphate isomerase) step.Source: proposal — no TS observed; Glu187-deprotonated water attacking the imine carbon + Lys229 regeneration per M-CSA Entry 222 (Base-assignment card)
Sequence DNA/gene NCBI Gene 226 (ALDOA) · protein UniProt P04075 (human) · rabbit P00883. Residue numbers on screen follow the PDB-author numbering of the rabbit structures (1ZAI/6ALD/1ADO chain A), i.e. UniProt P00883 − 1: Lys229 = Lys230 (UniProt/M-CSA), Asp33 = Asp34, Glu187 = Glu188.

How to tell a proposal from a fact on screen PROPOSAL markers (amber chip, clickable) + amber dotted phrases = mechanism inferred from crystal geometry and kinetics, not directly measured: the Asp33 general base (③), the Glu187-activated water path (④), and the schematic imine-frame offset of the 6ALD Michaelis pose (①, bottom of barrel). Statements without a marker = deposited geometry (e.g. the 1ZAI imine with C2–NZ = 1.30 Å, or the deposited 6ALD C2…NZ ≈ 7.3 Å). A transition state is never observed, so every arrow here is a literature-based proposal.

Legend & Fidelity

Mechanism fidelity — measured vs. inferred The measured layer: the rabbit backbone (1ZAI/6ALD/1ADO), the trapped imine geometry (C2–NZ re-derived and re-checked against the deposit, Rule 34 geomClaims), the 6ALD substrate pose, and the seated C1-phosphate. The inferred layer: the imine-frame offset that re-measures the same Michaelis pose at ≈ 6.83 Å (vs. the 6ALD-frame 7.27 Å) and the drawn closing of that gap to the attack line, the Asp33 proton abstraction that opens C3–C4, and the Glu187-activated water that hydrolyzes the imine. No retro-aldol transition state is observed, so both chemistry steps are literature-based proposals (M-CSA Entry 222).
Why is the Michaelis pose re-anchored "[schematic]"? Because the trapped 6ALD substrate is a non-covalent, Michaelis-like complex whose C2 carbonyl sits ≈ 7.3 Å from Lys229-NZ in the 6ALD frame — not at attack distance. "Re-anchor" here is a frame offset: the same deposited pose, shown in the 1ZAI imine frame, re-reads at ≈ 6.83 Å (the two lysine seats differ between the crystals; the substrate is not rotated). Closing that residual gap to the ≈ 3 Å of C–N bond formation is the literature-based adjustment, drawn schematically. The deposited substrate coordinates and the 1ZAI imine are measured; the bridge drawn between them is reconstruction.
Base assignment — Asp33 and Glu187, complementary not competing Asp33 can be seen as the general base whose proton relay (C3) opens the occasional debate about proton chops — we follow M-CSA Entry 222: Asp33 abstracts toward C3–C4 cleavage while Glu187 handles the dehydration (imine formation) and the hydrolysis water — the two residues are complementary, not competing assignments. Product order (G3P before DHAP) follows the M-CSA scheme.
Precision vs. accuracy — Å numbers are single crystallographic states Distances shown are single values from the deposited entries (1ZAI imine: C2–NZ 1.30 Å) or from our own common-frame superposition (6ALD Michaelis: 7.27 Å deposited, 6.83 Å for the same pose read in the imine frame; Ser271-OG…P1 3.8 Å) — not measured dynamics along the trajectory. This scene joins two structures (1ZAI imine frame + 6ALD substrate) in one frame; inter-state transitions are stitched and schematic. The gold H⁺ ball and proton arrow are deliberately drawn cartoons (Rule 23) illustrating the path, not a quantum / H-transfer trajectory. Treat every Å as a plausible crystallographic value, not a measured coordinate.
Physiological regulation — ALDOA in context Class-I aldolase A is regulated chiefly by gene expression and tissue isozyme distribution (ALDOA muscle/fetal tissues, ALDOB liver/kidney/intestine = hereditary fructose intolerance, ALDOC brain) rather than by a potent allosteric feedback brake — unlike, e.g., HK1. The reaction is freely reversible: in glycolysis the cleavage direction (F1,6BP → DHAP + G3P) dominates, while gluconeogenesis (e.g. in liver) runs the aldol condensation direction. DHAP and G3P are rapidly interconverted by triose phosphate isomerase, pulling the aldolase equilibrium forward.
Kinetics scope — one event, not a timecourse This tool animates a single catalytic event; it does not model timecourses, steady-state kinetics, or K_m / k_cat / V_max · [S] behavior. For stoichiometric context, BRENDA EC 4.1.2.13 tabulates human FBP-aldolases — Aldolase B (liver, wild-type) Km(D-fructose-1,6-BP) ≈ 0.004 mM at 22 °C / pH 7.6 (ref 651985 = Rellos P, Sygusch J & Cox TM, J Biol Chem 275:1145–1151, 2000), with the human isoform window 0.0017–0.052 mM across refs 4925 (erythrocyte ALDOA, Yeltman & Harris 1982) / 649476 (ALDOB, Malay/Procious/Tolan 2002) / 663932 (ALDOA, Esposito et al. 2004) — the same order as the rabbit muscle ALDOA animated here.
Sources — bibliography of this page S1 — PDB 1ZAI: rabbit aldolase A trapped imine (Schiff-base) complex — C2–NZ measured, contact network Lys146 / Glu187 / Asp33 to the C4-hydroxy (values on the Mechanism fidelity card); St-Jean et al. 2005, PMID:15870069 S2 — PDB 6ALD: non-covalent Michaelis-like complex with the substrate pose — the C2 carbonyl⋯Lys-NZ separation as deposited, and the same pose re-read in the 1ZAI imine frame (values on the Mechanism fidelity card; a frame offset, not a re-fit) S3 — PDB 1ADO: third rabbit ALDOA frame (backbone set 1ZAI/6ALD/1ADO chain A, PDB-author numbering = UniProt P00883 − 1) S4 — M-CSA entry 222: Asp33 general base abstracting the C3 proton as C3–C4 breaks, G3P-leaves-first order; role table lists Tyr363 (C3 pro-S removal), Ser300 (enamine stabilizer), Glu189 (water activation); DHAP–enamine reference PDB 2QUT S5 — St-Jean et al. 2005, PMID:15870069: Glu187-route alternative for C3–C4 cleavage (complementary, not competing, with M-CSA Asp33) S6 — Kinetics context: BRENDA EC 4.1.2.13 — Aldolase B (liver WT) K_m(FBP) and the human isoform window values quoted in the EvidenceView body, ref 651985 (Rellos, Sygusch & Cox, J Biol Chem 275:1145–1151, 2000); human isoform window refs 4925 (Yeltman & Harris 1982) · 649476 (Malay/Procious/Tolan 2002) · 663932 (Esposito et al. 2004) S7 — UniProt P04075 (human ALDOA) · rabbit P00883 · NCBI Gene 226 (ALDOA); isozymes ALDOA/ALDOB/ALDOC (ALDOB = hereditary fructose intolerance) S8 — Repo display conventions: Rule 30 (narrated claims reconstructible from the scene) · Rule 34 geometry (14 geomClaims in tools/manifests/ald.json, re-measured from the deposits) · every Å = a single crystallographic value (Precision vs. accuracy card) · [schematic] = frame offset / literature-based bridge between measured states, no deposition claimed; no retro-aldol TS is ever observed

› Backbones and ligand poses are taken directly from the listed PDB entries, superposed into one common frame. Where no crystalline state exists for a step (e.g., Michaelis geometry), the pose is drawn schematically using deposited geometries (6ALD substrate) and is labeled as such in the narration. Residue numbers follow the PDB-author numbering of the rabbit structures (1ZAI/6ALD/1ADO chain A) = UniProt P00883 − 1; human ALDOA (P04075) is homologous. Where inter-state transitions are stitched, the recomposed coordinates are schematic — see "Precision vs. accuracy" above.

N-barrel halfC-barrel halfONPMg/K