Malate Dehydrogenase - 3D Mechanism Simulator

Loading structural data… (2DFD)

References & Databases

START HERE — how to read L-malate + NAD⁺ → OAA + NADH + H⁺ A single oxidation cartoon on the human mitochondrial MDH2 pocket. Real crystal: 2DFD (1.9 Å), a ternary of L-malate (LMR) — the actual substrate, not an analog — and NAD⁺. His182 ★ is the general base (NE2 ⋯ O2 = 2.58 Å). Hydride donor is malate C2 (not C3) to nicotinamide C4 (C2 ⋯ C4N = 3.03 Å). NAD⁺ is staged first (compiled ordered-bi-bi; not a kinetic fit). Gold H⁺ parks on His182; blue H⁻ is the hydride. Oxaloacetate is [schematic]. M-CSA 526 writes proton+hydride as one catalog step; this page splits them. Pig His186–Asp158 maps here as His182; Asp155 is named, not drawn.
Crystal structure PDB 2DFD — human mitochondrial MDH2, 1.9 Å, Structural Genomics Consortium. Coordinates are public. Ligands: LMR ((2S)-2-hydroxybutanedioate = L-malate, the real substrate) and NAD⁺. Not ligand-free. Not an analog co-crystal.
Mechanism — His general base, hydride to C4 M-CSA 526 (pig cytoplasmic MDH, PDB 4MDH) writes a His186–Asp158 dyad: His abstracts the C2-OH proton; a hydride goes to NAD C4; Asp158 electrostatically stabilises the imidazolium. Primary structures: Birktoft et al., J Biol Chem 1983, PMID:6848515 · Birktoft et al., Biochemistry 1989, PMID:2775751. Human 2DFD map: catalytic His → His182 ★. Nearest Asp on this slice = Asp155 (CA⋯His182 CA, not a dyad claim) — named, not drawn. Chapman 1999 (PMID:10075524) is a pig cMDH specificity ternary (α-ketomalonate + tetrahydroNAD), not the kinetic or His-base source. Source: M-CSA 526 · Birktoft 1983 · Birktoft 1989 · Chapman 1999 (specificity only)
0 — Setup: the ternary pocket, shown empty 2DFD holds L-malate + NAD⁺. The pocket is shown empty so ordered binding can start. Opaque, run:null (no autoplay). His182 ★ is the general base. Asp155 is named, not drawn.Source: PDB 2DFD
① — NAD⁺ binds (ordered, first) Crystal NAD⁺ docks into the Rossmann fold. Nicotinamide C4 is the hydride acceptor. NAD-first order is compiled MDH kinetic literature, not a fit on this page and not Chapman 1999 (that paper is a specificity co-crystal).Source: 2DFD NAD pose · compiled ordered-bi-bi kinetics (not re-fit)
② — L-Malate binds (2-OH on His182) Crystal LMR — the real substrate — docks. O2 ⋯ His182 NE2 = 2.58 Å (measured on this ternary). Arg86 / Arg92 / Arg158 grip the dicarboxylate.Source: 2DFD LMR atoms
③ — His182 ★ abstracts the 2-OH proton (hypothetical, lit.-based) A gold H⁺ sphere rides a gold arrow from LMR O2 to His182 NE2 and parks. The alkoxide label is a teaching overlay. M-CSA 526 writes this proton transfer in the same step as the hydride; this page splits them. Asp155 is named, not drawn.Source: M-CSA 526 step 1 (concerted on the catalog) · Birktoft 1983/1989
④ — Hydride C2 → NAD⁺ C4 → OAA + NADH (hypothetical, lit.-based) Blue H⁻ (hydride) rides from malate C2 to NAD C4N (3.03 Å). L-malate fades in place to schematic oxaloacetate; NAD⁺ relabels NADH. Parked gold H⁺ stays on His182. A-side / pro-R nicotinamide chemistry is compiled 2-hydroxyacid DH literature, not re-derived from Chapman 1999. Slow-motion on purpose.Source: 2DFD C2⋯C4N · M-CSA 526 hydride to C4 · compiled A-side/pro-R
⑤ — H⁺ released — His182 regenerates (hypothetical, lit.-based) The parked gold H⁺ leaves to solvent. M-CSA 526 step 2 marks His deprotonation as an inferred regeneration. Net stoichiometry still needs that H⁺ (EC 1.1.1.37).Source: M-CSA 526 step 2 (inferred) · EC 1.1.1.37 net H⁺
⑥ — OAA + NADH leave — cycle closes [schematic] Oxaloacetate and NADH depart in opposite directions. OAA re-enters citrate synthase (TCA step 1); NADH feeds the ETC.Source: TCA cycle bookkeeping

How to read this page — a first-pass primer First pass: Setup (empty pocket, His182 ★) → ① NAD⁺ docks → ② L-malate docks, 2-OH on His182 → ③ gold H⁺ O2 → NE2 (parks) → ④ blue hydride C2 → C4N, malate becomes OAA, NAD⁺ becomes NADH → ⑤ gold H⁺ leaves → ⑥ OAA and NADH leave opposite ways. Second pass: 2.58 Å and 3.03 Å are crystal facts on this ternary; OAA and the H spheres are [schematic]. Third pass: open “Disagreements” for concerted vs split, and M-CSA 526 pig numbering vs 2DFD His182.
Chemistry primer — NAD⁺-dependent oxidation L-malate is a C4 hydroxy-acid. Removing the 2-OH proton and the C2 hydride leaves a ketone at C2 = oxaloacetate. NAD⁺ accepts that hydride at nicotinamide C4 (compiled A-side / pro-R). Net H⁺ is released to solvent. Gold = H⁺. Blue = hydride H⁻ (hydrogen plus two electrons). The reaction is reversible; this page draws the TCA oxidative direction.
Words used on screen — ordered bi-bi · alkoxide · pro-R Ordered bi-bi — NAD⁺ binds before L-malate in compiled MDH kinetics; this page is staging, not a fit. Alkoxide — C2–O⁻ after His takes the 2-OH proton; a teaching label, not a deposited species. pro-R / A-side — compiled 2-hydroxyacid DH nicotinamide face, not a number from 2DFD. LMR — PDB ligand code for L-malate on 2DFD, not an analog.

Legend & Fidelity

Mechanism fidelity — measured vs. inferred The measured layer: 2DFD (1.9 Å) ternary with LMR + NAD⁺; His182 NE2 ⋯ LMR O2 = 2.58 Å; LMR C2 ⋯ NAD C4N = 3.03 Å. The inferred layer ([schematic]): oxaloacetate body, transferring H⁺/H⁻ spheres, NAD⁺→NADH label switch, product leave, and ⑤ His regeneration (M-CSA 526 step 2). Binding order is compiled staging, not a kinetic fit. The protein mesh is static.
Disagreements on record (a) Concerted vs split. M-CSA 526 step 1 writes His proton abstraction and the hydride to NAD C4 as one step. This page splits them (③ then ④) for 1-event/1-step teaching — the catalog is concerted, not an opposing minority view. (b) M-CSA numbering. Entry 526 is pig cytoplasmic His186–Asp158 (4MDH). This page uses 2DFD His182 ★. Asp155 is the nearest Asp by CA on this slice — named, not drawn — not asserted as the catalog Asp. (c) Chapman 1999. Pig cMDH · α-ketomalonate · tetrahydroNAD specificity ternary — not the His-base, ordered-bi-bi, or pro-R primary source. (d) Isozyme. MDH2 is mitochondrial; MDH1 (cytosolic) is another gene, not this mesh. (e) Direction. Fully reversible; the page draws the TCA oxidative direction. (f) ⑤ regeneration. M-CSA step 2 is inferred; the screen flags it.
Regulation — OAA product · isoforms Oxaloacetate is both the product here and a tight inhibitor of upstream SDH — named, not drawn as an MDH·OAA crystal. Mitochondrial MDH2 (this page) vs cytosolic MDH1 share the His/NAD chemistry but are different genes. No allosteric effector is asserted from this 2DFD frame.Source: textbook MDH isoforms · SDH/OAA inhibition (named, not simulated)
Kinetics scope (no measured timecourse on this page) One L-malate → OAA + NADH + H⁺ event is drawn, not a steady-state run. Compiled BRENDA values for EC 1.1.1.37 are not re-asserted here — no measured k_cat/K_m on this page [schematic]. Pair with a kinetics chapter.Source: BRENDA EC 1.1.1.37 — no k_cat/K_m re-asserted
Sequence Human MDH2 (mitochondrial) UniProt P40926 · NCBI Gene 4191. Screen uses 2DFD PDB-author numbering (His182 = mature chain; UniProt precursor is 338 AA).
Simulation map — what you see is what is written Setup = 2DFD pocket empty (opaque, run:null). ① NAD⁺ moves in (crystal). ② L-malate moves in (crystal); 2.58 Å label after dock. ③ gold H⁺ + gold arrow O2 → His182 NE2 (parks). ④ slow-mo: blue hydride H⁻ + blue arrow C2 → C4N; LMR fades to OAA; NAD⁺ relabels NADH; parked gold H⁺ remains. ⑤ gold H⁺ leaves to solvent (inferred regeneration). ⑥ OAA and NADH leave opposite ways. Only these events move.
Sources — bibliography of this page S1 — PDB 2DFD: human mitochondrial MDH2, 1.9 Å, ternary L-malate (LMR) + NAD⁺ (SGC) — public coordinates; measured NE2⋯O2 = 2.58 Å and C2⋯C4N = 3.03 Å S2 — Mechanism: M-CSA 526 · Birktoft et al., J Biol Chem 1983, PMID:6848515 · Birktoft et al., Biochemistry 1989, PMID:2775751 — His–Asp dyad; hydride to nicotinamide C4. Step 1 is concerted on the catalog; step 2 His regeneration is inferred S3 — Chapman AD et al., J Mol Biol 1999, PMID:10075524 — pig cytoplasmic MDH specificity ternary (α-ketomalonate + tetrahydroNAD). Not the ordered-bi-bi or His-base primary source. Human map His186→His182; Asp155 named, not drawn S4 — Sequence: UniProt P40926 (human MDH2) · NCBI Gene 4191 S5 — Kinetics: BRENDA EC 1.1.1.37 — 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 identity, H⁺/H⁻ flight, product release), the pose is drawn schematically and labeled as such in the narration.

NAD-binding domainC-domainONH⁺ (gold)H⁻ hydride (blue)