- START HERE — how to read 2 acetyl-CoA → acetoacetyl-CoA + CoA A thiolase cartoon on the human ACAT1 / T2 frame that actually exists: 2IBY (1.85 Å X-ray) chain A with crystal free CoA + K+ + Cl−. Occupancy is free CoA, not acetyl-CoA, not acetoacetyl-CoA — never read CoA as acetyl-CoA. Cys126 on this crystal is CSO (S-hydroxycysteine), not acetyl-Cys. Teaching Cys126 omits the extra OD. Cys126 ★ is the nucleophilic cysteine (UniProt ACT_SITE). His385 is named, not starred (UniProt Site 385). Cys413 is named — UniProt P24752 ACT_SITE is Cys413, not His413. Covalent E~acetyl is compiled and disclosed — not a pure A-page. K+ stays. Blue is nucleophile, not hydride. V34A is the construct mutation; residue 34 is not on this mesh. ACAT2 and ACAA2 are other genes. Measured Å are deposited 2IBY free-CoA pairs only.
- Crystal / X-ray structures PDB 2IBY — human mitochondrial acetoacetyl-CoA thiolase + CoA + K + Cl, 1.85 Å X-ray (Haapalainen 2007). Coordinates are public. Occupancy is free CoA. Cys126 is CSO. V34A is engineered. Human ACAT1 UniProt P24752.
- Mechanism ΓÇö Cys126 nucleophile; condensation direction UniProt P24752 EC 2.3.1.9: 2 acetyl-CoA = acetoacetyl-CoA + CoA. This page teaches condensation (ketogenesis). Thiolysis of acetoacetyl-CoA is ACAA2ΓÇÖs ╬▓-ox page, not this gene. On this mesh the nucleophilic cysteine is Cys126. His385 is named, not starred. Covalent acetyl-Cys126 is compiled, not this occupancy. ACAT2 is another gene. Source: UniProt P24752 ┬╖ Haapalainen 2007
- 0 — Setup: empty thiolase pocket 2IBY occupancy is free CoA + K + Cl, not acetyl-CoA, not acetoacetyl-CoA. Cys126 is CSO, not acetyl-Cys. The pocket is shown empty so binding can start. K+ stays. Opaque, run:null (no autoplay). Cys126 ★ is the nucleophile. V34A is the construct mutation; residue 34 is not on this mesh (Cα 37–427). MES / GOL / Cl are named, not substrate. ACAT2 and ACAA2 are other genes.Source: PDB 2IBY (1.85 Å X-ray)
- ① — First acetyl-CoA binds [schematic] Acetyl-CoA docks on the 2IBY CoA pose — not a deposited acetyl-CoA. CoA on 2IBY is free CoA, not acetyl-CoA. Overlay Å are not geomClaims. K+ stays.Source: 2IBY CoA pose + 1F0Y CAA thiol-end overlay
- ② — Cys126 attacks carbonyl → acetyl-enzyme + CoA (hypothetical, lit.-based) Blue nucleophile rides from Cys126 SG onto schematic acetyl C1. Acetyl-CoA cross-fades to crystal CoA and a compiled acetyl-Cys126 stub. Covalent E~acetyl is disclosed — not a pure A-page. This crystal has CSO, not acetyl-Cys. Slow-motion on purpose. Blue = nucleophile, not hydride.Source: compiled thiolase ping-pong · UniProt P24752 Cys126
- Γæó ΓÇö First CoA leaves Crystal CoA departs. Compiled acetyl-Cys126 stays. K+ stays. Cys126 is CSO on 2IBY, not acetyl-Cys. Free CoA is not acetyl-CoA.Source: thiolase ping-pong bookkeeping
- ④ — Second acetyl-CoA binds [schematic] Second acetyl-CoA docks on the 2IBY CoA pose. Cys126 SG ⋯ CoA S1P = 4.26 Å is the deposited free-CoA contact, not an acetyl-enzyme geometry.Source: PDB 2IBY CoA
- ⑤ — Acetyl-Cys C2 attacks → acetoacetyl-CoA (hypothetical, lit.-based) Blue nucleophile rides from compiled acetyl-Cys C2 onto the second acetyl carbonyl. Second acetyl-CoA + acetyl stub cross-fade to schematic acetoacetyl-CoA. His385 is named, not starred; Cys413 is named. Cys126 SG ⋯ His385 NE2 = 3.45 Å is deposited 2IBY. Slow-motion on purpose. Blue = nucleophile, not hydride.Source: compiled thiolase condensation · 2IBY Cys126 / His385
- ⑥ — Acetoacetyl-CoA leaves — HMGCS2 next Schematic acetoacetyl-CoA departs. K+ stays. This page does not run ACAA2 thiolysis or ACAT2. Cys126 is CSO on 2IBY, not acetyl-Cys. Overlay Å are not geomClaims. K+ ⋯ Tyr219 OH = 2.74 Å is deposited 2IBY.Source: ketogenesis bookkeeping · EC 2.3.1.9
- How to read this page — a first-pass primer First pass: Setup (empty pocket, Cys126 ★, CSO not acetyl, K stays, V34A) → ① schematic first acetyl-CoA docks on CoA pose → ② blue Cys126-S → carbonyl, fade to CoA + acetyl-Cys → ③ CoA leaves → ④ schematic second acetyl-CoA docks → ⑤ acetyl-Cys C2 → carbonyl, fade to acetoacetyl-CoA → ⑥ leave; HMGCS2 next. Second pass: 4.26 / 3.45 / 2.74 Å are crystal facts on 2IBY free-CoA + K complex; acetyl-CoA and acetyl-Cys are [schematic]; CoA is free CoA, not substrate. Third pass: open “Disagreements” for CSO vs Cys, free CoA vs acetyl, UniProt P24752 Cys413, not His413, and ACAT2 / ACAA2 vs this gene.
- Chemistry primer ΓÇö ping-pong thiolase (condensation) ACAT1 Cys126 attacks the first acetyl-CoA carbonyl, releasing CoA and leaving an acetyl-enzyme. A second acetyl-CoA then condenses with that acetyl-Cys to acetoacetyl-CoA. Blue is nucleophile, not hydride. This is not ACAA2 (EC 2.3.1.16 thiolysis) and not ACAT2 (cytosolic).
- Words used on screen ΓÇö free CoA ┬╖ CSO ┬╖ Cys126 Γÿà ┬╖ compiled acetyl-Cys ┬╖ K stays Free CoA ΓÇö 2IBY occupancy, not acetyl-CoA, not acetoacetyl-CoA. CSO ΓÇö Cys126 is S-hydroxycysteine on this crystal; teaching omits OD. Cys126 Γÿà ΓÇö nucleophile on this PDB = UniProt. Compiled acetyl-Cys ΓÇö covalent intermediate, not this occupancy. K stays ΓÇö deposited potassium, activator, not a reactant.
Legend & Fidelity
- Mechanism fidelity — measured vs. inferred The measured layer: 2IBY (1.85 Å X-ray) human ACAT1 + free CoA + K + Cl; Cys126 SG ⋯ CoA S1P = 4.26 Å; Cys126 SG ⋯ His385 NE2 = 3.45 Å; K+ ⋯ Tyr219 OH = 2.74 Å. The inferred layer ([schematic]): acetyl-CoA on the CoA pose, transferring nucleophile spheres, acetyl-Cys stub, acetoacetyl-CoA product. Binding order is compiled staging, not a kinetic fit. The protein mesh is static. Free-CoA Å are not acetyl-CoA geomClaims. Cys126 is CSO, not acetyl-Cys.
- Disagreements on record (a) Occupancy vs substrate. 2IBY holds free CoA, not acetyl-CoA, not acetoacetyl-CoA. (b) CSO vs Cys. Cys126 is S-hydroxycysteine on this crystal ΓÇö not acetyl-Cys and not unmodified Cys. Teaching omits OD. (c) Covalent acetyl-Cys126 is compiled identity, not this occupancy. (d) His385 is named, not starred. (e) UniProt P24752 ACT_SITE is Cys413, not His413 ΓÇö labels use PDB ids. (f) Concerted vs split nucleophile/condensation ΓÇö this page splits Γæí then Γæñ for 1-event/1-step teaching. (g) ACAT2 is cytosolic, another gene. (h) ACAA2 is T1 thiolysis (EC 2.3.1.16), another gene. (i) V34A is engineered; residue 34 is not on this mesh (C╬▒ 37ΓÇô427). (j) MES / GOL / Cl are named, not substrate. (k) RHEA:30719 methyl-branch activity is not shown. (l) OXCT1 is ketolysis, not this path.
- Regulation ΓÇö K+ ┬╖ isoforms ┬╖ tetramer K+ activates (not Na+) ΓÇö Haapalainen 2007. This cartoon does not simulate ion-binding kinetics. This condensation is monofunctional ACAT1 / T2 only. Cytosolic ACAT2 is another gene ΓÇö not this mesh. ACAA2 / T1 is ╬▓-ox thiolysis, another gene. Homotetramer assembly is not this cartoon. Matrix localization is assumed, not a catalytic event on this page.Source: UniProt P24752 ┬╖ PDB 2IBY ┬╖ Haapalainen 2007
- Kinetics scope (no measured timecourse on this page) One 2 acetyl-CoA → acetoacetyl-CoA + CoA event is drawn, not a steady-state run. Compiled BRENDA values for EC 2.3.1.9 are not re-asserted here — no measured kcat/Km on this page [schematic]. Pair with a kinetics chapter.Source: BRENDA EC 2.3.1.9 — no kcat/Km re-asserted
- Sequence Human ACAT1 UniProt P24752 ┬╖ NCBI Gene 38. This screen uses 2IBY numbering (Cys126 = UniProt Cys126). Chain A. C╬▒ 37ΓÇô427. Transit 1ΓÇô33 and V34A are not this mesh.
- Simulation map — what you see is what is written Setup = 2IBY pocket empty, K stays (opaque, no autoplay). ① schematic first acetyl-CoA moves in. ② slow-mo: blue Cys126-S → carbonyl; fade to CoA + acetyl-Cys. ③ CoA leaves. ④ schematic second acetyl-CoA moves in. ⑤ slow-mo: acetyl-Cys C2 → carbonyl; fade to acetoacetyl-CoA. ⑥ acetoacetyl-CoA leaves. Only these events move.
- Sources — bibliography of this page S1 — PDB 2IBY: human ACAT1 + CoA + K + Cl, 1.85 Å X-ray — public coordinates; measured Cys126 SG⋯CoA S1P = 4.26 Å · Cys126 SG⋯His385 NE2 = 3.45 Å · K+⋯Tyr219 OH = 2.74 Å S2 — Haapalainen et al., 2007, PMID:17371050 — human mitochondrial acetoacetyl-CoA thiolase; K+ and Cl− S3 — Sequence: UniProt P24752 · NCBI Gene 38 S4 — Kinetics: BRENDA EC 2.3.1.9 — no kcat/Km re-asserted on this page
› Backbones and CoA poses are taken directly from the listed PDB entries, superposed into one common frame. Where no crystalline state exists for a step (e.g., acetyl-CoA on 2IBY, acetyl-Cys126, nucleophile flight, product leave), the pose is drawn schematically and labeled as such in the narration. Occupancy is free CoA, not acetyl-CoA. Cys126 is CSO, not acetyl-Cys. Free-CoA Å are not acetyl-CoA geomClaims.