Research ArticleSTRUCTURAL BIOLOGY

High-resolution cryo-EM structures of respiratory complex I: Mechanism, assembly, and disease

See allHide authors and affiliations

Science Advances  11 Dec 2019:
Vol. 5, no. 12, eaax9484
DOI: 10.1126/sciadv.aax9484
  • Fig. 1 Cryo-EM structure of respiratory complex I from Y. lipolytica.

    (A) Side view; inset shows FMN and FeS clusters [assignment of EPR signatures N1 to N5 according to (72)] and a Q molecule in the access pathway. (B) Top view with the matrix arm omitted for clarity and lipid molecules (yellow) and Q molecule (green) in sphere representation.

  • Fig. 2 Q molecule in the access pathway.

    (A) Slice of membrane arm/matrix arm interface viewed from the back of complex I, a cavity (gray) opens in ND1 and permits access of Q from the bilayer to the Q reduction site near cluster N2. (B) Selected residues near the Q binding site (density map drawn at 1.0 σ contour level in blue mesh; Q molecule, green sticks) (see fig. S4).

  • Fig. 3 Lipid binding sites and unique protein lipid arrangement at the junction of the membrane and matrix arms.

    (A) Lipid and detergent binding sites in the membrane arm. CL, cardiolipin; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PI, phosphatidylinositol. (B) Cross section of a simulation snapshot reveals that the lipid bilayer thickness is reduced by ~10 Å in the vicinity of protein, apparently due to the unusual local protein architecture. Q is shown as spheres colored by atom (carbon, green; oxygen, red). Lipids in the two leaflets are colored by lipid type. PC/PE/CDL in the matrix leaflet are shown in shades of pink, and those in the other leaflet in shades of blue. Highly bent lipid tails next to protein are drawn as sticks. Subunits are colored as in Fig. 1, and accessory subunits are gray.

  • Fig. 4 Mutant lacking NDUFS4.

    (A) Overlay of wild type (gray, NDUFS4 green surface representation) and mutant (cartoon, color as in Fig. 1). (B) Zoom into membrane arm. Disordered protein in the mutant is highlighted by color. (C) Overlay of NDUFS4 on the matrix arm of the mutant. (D) FeS clusters N1b and N3 in the mutant are more exposed to solvent than in the fully assembled complex (see main text).

  • Fig. 5 Cryo-EM structure of a complex I assembly intermediate.

    (A) Overlay of wild type (gray) and assembly intermediate (color) with assembly factor NDUFAF2 (purple surface). (B) Overlay as in (A) viewed from the back. The arrow indicates the tilted matrix arm. (C) Membrane arm/matrix arm interface of wild-type complex I viewed as in (B) with NDUFA12 and NDUFS6 highlighted. (D) Assembly intermediate with NDUFAF2 highlighted, viewed as in (B); compare fig. S13.

  • Fig. 6 Function of the assembly factor NDUFAF2.

    (A) Assembly factor NDUFAF2 associates with a complex I assembly intermediate that lacks the N module (NDUFS1, NDUFV1, and NDUFV2). NDUFAF2 provides a platform for N module attachment (dashed line) and anchors NDUFS1 with its C-terminal tail. In the wild type, the concerted interplay of NDUFS4, NDUFS6, and NDUFA12 releases NDUFAF2. NDUFS6 and NDUFA12 form a structural unit that is obligatory for function. (B) In the NDUFS6Δ mutant, NDUFS4 can bind and then detaches a part of NDUFAF2 (dotted line), but the assembly factor remains firmly bound because NDUFS6 is absent.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/12/eaax9484/DC1

    Supplementary Materials and Methods

    Fig. S1. Structure determination of wild-type complex I, assembly intermediate, and mutant without NDUFS4.

    Fig. S2. Representative cryo-EM densities of wild-type complex I with fitted models.

    Fig. S3. Assignment of accessory subunits.

    Fig. S4. A second Q binding site in complex I and quantitative mass spectrometry.

    Fig. S5. Atomic model of Y. lipolytica complex I in membrane-solvent environment and hydrogen bond analysis from MD simulation trajectory.

    Fig. S6. Interactions between protein and the Q head group and tail.

    Fig. S7. Lipid binding sites in yeast and mammalian complex I.

    Fig. S8. Surface helices and lipids modeled at the interface of the membrane arm and the peripheral arm.

    Fig. S9. Bending of the lipid tails seen in MD simulation.

    Fig. S10. Spectrophotometric assay of NADH:DBQ activity.

    Fig. S11. Conformation of the β1-β2 loop of NDUFS2 and loops and regions critical for the active/deactive transition.

    Fig. S12. Proton pumping activity of the NDUFS4 mutant.

    Fig. S13. Cartoon representation and sequence alignment of NDUFAF2 and NDUFA12 from Y. lipolytica and Homo sapiens.

    Table S1. Data collection, refinement, and model statistics.

    Table S2. Subunit names and chain identifiers for Y. lipolytica and human and bovine complex I.

    Table S3. Lipid species detected in Y. lipolytica complex I.

    Movie S1. Cryo-EM structure of respiratory complex I from Y. lipolytica in LMNG at 3.2-Å resolution.

    Movie S2. Cryo-EM structure of ΔNDUFS4 at 4.0-Å resolution.

    Movie S3. Cryo-EM structure of the ΔNDUFS6 assembly intermediate at 3.3-Å resolution.

    References (7378)

  • Supplementary Materials

    The PDFset includes:

    • Supplementary Materials and Methods
    • Fig. S1. Structure determination of wild-type complex I, assembly intermediate, and mutant without NDUFS4.
    • Fig. S2. Representative cryo-EM densities of wild-type complex I with fitted models.
    • Fig. S3. Assignment of accessory subunits.
    • Fig. S4. A second Q binding site in complex I and quantitative mass spectrometry.
    • Fig. S5. Atomic model of Y. lipolytica complex I in membrane-solvent environment and hydrogen bond analysis from MD simulation trajectory.
    • Fig. S6. Interactions between protein and the Q head group and tail.
    • Fig. S7. Lipid binding sites in yeast and mammalian complex I.
    • Fig. S8. Surface helices and lipids modeled at the interface of the membrane arm and the peripheral arm.
    • Fig. S9. Bending of the lipid tails seen in MD simulation.
    • Fig. S10. Spectrophotometric assay of NADH:DBQ activity.
    • Fig. S11. Conformation of the β1-β2 loop of NDUFS2 and loops and regions critical for the active/deactive transition.
    • Fig. S12. Proton pumping activity of the NDUFS4 mutant.
    • Fig. S13. Cartoon representation and sequence alignment of NDUFAF2 and NDUFA12 from Y. lipolytica and Homo sapiens.
    • Table S1. Data collection, refinement, and model statistics.
    • Table S2. Subunit names and chain identifiers for Y. lipolytica and human and bovine complex I.
    • Table S3. Lipid species detected in Y. lipolytica complex I.
    • Legends for movies S1 and S3
    • References (7378)

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Cryo-EM structure of respiratory complex I from Y. lipolytica in LMNG at 3.2-Å resolution.
    • Movie S2 (.mp4 format). Movie S2. Cryo-EM structure of ΔNDUFS4 at 4.0-Å resolution.
    • Movie S3 (.mp4 format). Movie S3. Cryo-EM structure of the ΔNDUFS6 assembly intermediate at 3.3-Å resolution.

    Files in this Data Supplement:

Stay Connected to Science Advances

Navigate This Article