PEP Carboxykinase - 3D Mechanism Simulator

Loading structural data… (1KHB)

References & Databases

START HERE — how to read OAA + GTP analog → PEP + CO₂ + GDP A decarboxylation-then-phosphoryl cartoon on the human cytosolic PEPCK frame that actually exists: 1KHB (1.85 Å) with a nonhydrolyzable GCP analog and two Mn²⁺. GCP is phosphomethylphosphonic guanylate ester — not GTP. OAA, enolpyruvate, CO₂, and GDP are [schematic]. PEP is a 1KHF overlay (Cα RMSD 0.279 Å), not a ternary. Cys288 is the UniProt P-loop marker, not a general base. Compiled PEPCK writes decarboxylation then phosphoryl transfer; this page splits them. This is PCK1 (cytosol), not mitochondrial PCK2.
Crystal structures PDB 1KHB — human cytosolic PEPCK, 1.85 Å, GCP + MN×2 (Dunten 2002). Coordinates are public. PDB 1KHF supplies the PEP overlay only. 1NHX is a xanthine-inhibitor crystal — wrong ligand, not used.
Mechanism — decarboxylation then phosphoryl transfer Dunten et al., J Mol Biol 2002, PMID:11851336 — human PCK1 with GCP. UniProt P35558 EC 4.1.1.32: OAA + GTP → PEP + CO₂ + GDP. Nucleotide Mn (MN1) holds GCP O3G / O2B; catalytic Mn (MN2) is the Lys244 / His264 / Asp311 shell. Overlay PEP O2⋯GCP PG is teaching geometry, not a geomClaim. Source: Dunten 2002 · UniProt P35558
0 — Setup: two Mn²⁺, pocket shown empty 1KHB holds GCP and two Mn. The pocket is shown empty so ordered binding can start. Both Mn ions stay as named occupants. GCP is a GTP analog — the β–γ bridge is a methylene. Opaque, run:null (no autoplay). Cys288 is P-loop, not a base.Source: PDB 1KHB
① — GTP analog binds (crystal) Crystal GCP docks. It cannot transfer phosphate on this crystal. Thr291 OG1 ⋯ MN1 = 2.22 Å. MN1 ⋯ O3G = 2.00 Å · MN1 ⋯ O2B = 2.17 Å. Binding order is compiled staging, not a fit.Source: 1KHB GCP A704 pose
② — Oxaloacetate binds (schematic) Schematic OAA is seated — not deposited on 1KHB. C2 is a ketone (not the PEP alkene). C3 is methylene; the C3 carboxyl is the leaving CO₂. 1NHX is not this ligand.Source: compiled PEPCK · 1KHB has no OAA
③ — OAA decarboxylates → enolpyruvate + CO₂ (hypothetical, lit.-based) OAA decarboxylates at C3. Schematic CO₂ is born at the C3 carboxyl; the remaining skeleton is enolpyruvate. No OAA or CO₂ is in 1KHB. Cys288 is not the base that takes this carbon.Source: compiled PEPCK decarboxylation · 1KHB occupancy
④ — CO₂ leaves (hypothetical, lit.-based) Schematic CO₂ departs. Enolpyruvate stays for the analog γ-phosphonate. Net EC 4.1.1.32 already counted this CO₂.Source: EC 4.1.1.32 bookkeeping
⑤ — Phosphoryl transfer → PEP (hypothetical, lit.-based) Enolpyruvate oxygen attacks analog PG. Enolpyruvate fades in place to PEP (1KHF overlay) and GCP to GDP. GCP cannot actually transfer — the methylene bridge is the analog hedge. Slow-motion on purpose.Source: compiled PEPCK phosphoryl · overlay 1KHF not ternary
⑥ — PEP leaves — FBPase is next PEP and schematic GDP depart in opposite directions. Both Mn²⁺ stay. PEP feeds the reversible stretch toward FBPase. This is cytosolic PCK1, not mitochondrial PCK2.Source: GNG bookkeeping

How to read this page — a first-pass primer First pass: Setup (two Mn present, pocket empty, GCP analog named) → ① GCP docks → ② schematic OAA docks → ③ decarboxylation, OAA becomes enolpyruvate, CO₂ born → ④ CO₂ leaves → ⑤ phosphoryl PG → enol O, PEP + GDP → ⑥ PEP and GDP leave. Second pass: 2.22 Å is a crystal fact on 1KHB; OAA, enolpyruvate, CO₂, GDP, and PEP overlay are [schematic]. Third pass: open “Disagreements” for analog vs GTP, overlay vs ternary, and Cys288.
Chemistry primer — decarboxylation then phosphoryl Cytosolic PEPCK decarboxylates oxaloacetate to an enolate, then transfers the GTP γ-phosphoryl onto that enolate to make PEP. This page uses a nonhydrolyzable analog (GCP) as the nucleotide stand-in. Two Mn²⁺: nucleotide metal (MN1) and catalytic metal (MN2). Gold is not used — the flying group here is a blue PO₃ cartoon.
Words used on screen — GCP · MN1 · MN2 · overlay PEP GCP — phosphomethylphosphonic guanylate ester (GTP analog; methylene β–γ bridge). MN1 — nucleotide Mn (residue 701). MN2 — catalytic Mn (residue 702). Overlay PEP — 1KHF PEP Kabsch-fit onto 1KHB, not a ternary crystal.

Legend & Fidelity

Mechanism fidelity — measured vs. inferred The measured layer: 1KHB (1.85 Å) with GCP + two Mn; Thr291 OG1 ⋯ MN1 = 2.22 Å; Mn shells 2.00–2.53 Å. The inferred layer ([schematic]): OAA, enolpyruvate, CO₂, GDP, PEP overlay, transferring PO₃ sphere, product leave. Binding order is compiled staging, not a kinetic fit. The protein mesh is static. Overlay contacts are not geomClaims.
Disagreements on record (a) GCP ≠ GTP. The methylene bridge cannot transfer on the crystal. (b) No OAA co-crystal. OAA / ENL / CO₂ / GDP are schematic. (c) PEP is overlay, not ternary. 1KHF onto 1KHB (RMSD 0.279 Å); PEP O2⋯GCP PG is not a geomClaim. (d) Cys288 is P-loop, not a base. UniProt marks 288; it does not abstract this carbon. (e) Concerted vs split. Compiled PEPCK writes decarboxylation then phosphoryl; this page splits CO₂ leave from transfer for 1-event/1-step teaching. (f) 1NHX is a xanthine inhibitor. (g) PCK2 is a different gene (mitochondrial).
Regulation — glucagon · cortisol · cytosolic vs mitochondrial PCK1 is the cytosolic gluconeogenic enzyme (this page). Mitochondrial PCK2 is a different gene — named, not this mesh. Transcriptional induction (glucagon / cortisol) is named, not simulated.Source: UniProt P35558 · Dunten 2002 PMID:11851336
Kinetics scope (no measured timecourse on this page) One OAA → PEP event is drawn, not a steady-state run. Compiled BRENDA values for EC 4.1.1.32 are not re-asserted here — no measured k_cat/K_m on this page [schematic]. Pair with a kinetics chapter.Source: BRENDA EC 4.1.1.32 — no k_cat/K_m re-asserted
Sequence Human PCK1 UniProt P35558 · NCBI Gene 5105. Screen uses 1KHB / UniProt numbering (Cys288, Lys244, Thr291). Cα 10–622; gaps 465–472 · 547–548 disordered.
Simulation map — what you see is what is written Setup = 1KHB pocket empty, two Mn²⁺ visible (opaque, run:null). ① GCP analog moves in (crystal). ② schematic OAA moves in. ③ OAA fades to enolpyruvate; schematic CO₂ is born. ④ CO₂ leaves. ⑤ slow-mo: PO₃ cartoon PG → enol O; enolpyruvate fades to PEP; GCP fades to GDP. ⑥ PEP and GDP leave opposite ways; both Mn stay. Only these events move.
Sources — bibliography of this page S1 — PDB 1KHB: human cytosolic PEPCK, 1.85 Å, GCP + MN×2 — public coordinates; measured OG1⋯MN1 = 2.22 Å S2 — PDB 1KHF: PEP overlay onto 1KHB (Cα RMSD 0.279 Å) — [schematic] product seat, not a ternary S3 — Dunten et al., J Mol Biol 2002, PMID:11851336 — human PCK1 + GCP S4 — Sequence: UniProt P35558 (human PCK1) · NCBI Gene 5105 S5 — Kinetics: BRENDA EC 4.1.1.32 — no k_cat/K_m re-asserted on this page

› 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., oxaloacetate, enolpyruvate, CO₂, GDP, PEP identity, PO₃ flight, product release), the pose is drawn schematically and labeled as such in the narration.

N-lobeC-lobeONPO₃ (blue)