Structural basis for electron transport mechanism of complex I-like photosynthetic NAD(P)H dehydrogenase

Abstract

NAD(P)H dehydrogenase-like (NDH) complex NDH-1L of cyanobacteria plays a crucial role in cyclic electron flow (CEF) around photosystem I and respiration processes. NDH-1L couples the electron transport from ferredoxin (Fd) to plastoquinone (PQ) and proton pumping from cytoplasm to the lumen that drives the ATP production. NDH-1L-dependent CEF increases the ATP/NADPH ratio, and is therefore pivotal for oxygenic phototrophs to function under stress. Here we report two structures of NDH-1L from Thermosynechococcus elongatus BP-1, in complex with one Fd and an endogenous PQ, respectively. Our structures represent the complete model of cyanobacterial NDH-1L, revealing the binding manner of NDH-1L with Fd and PQ, as well as the structural elements crucial for proper functioning of the NDH-1L complex. Together, our data provides deep insights into the electron transport from Fd to PQ, and its coupling with proton translocation in NDH-1L.

Fig. 1. The NDH-PQ and NDH-Fd structures.

a Cartoon representation of the NDH-PQ structure viewed along the membrane plane. PQ is shown as orange spheres and indicated. b The overall structure of NDH-Fd viewed along the membrane plane. c The location of OPS subunits in NDH-1L complex. The eight OPS subunits are colored differently and indicated, while other subunits are shown in white.

Fig. 2. The membrane arm with lipid and carotenoid molecules.

a Distribution of cofactors in NDH-Fd structure. Fe–S clusters are shown as spheres, PQ (orange), lipids (blue), and carotenoids (red) are shown as sticks. b Three lipid molecules (shown as ball-and-stick mode) contribute to the stabilization of NdhL, NdhA, NdhC, and NdhG. c The stromal side view of the lipid distribution at the distal part of the membrane arm. d Two carotenoid (BCR) and three lipid molecules participate in NdhF–NdhD interactions. e The binding pocket of BCR2. The color codes of subunits are the same as those in Fig. 1. The horizontal helix of NdhF is shown as blue cartoon. The BCR molecules are shown as red sticks. Lipid molecules are colored yellow for phosphatidyl glycerol (PG), gray for sulfoquinovosyldiacyl glycerol (SQDG), orange for monogalactosyldiacyl glycerol (MGDG), and blue for DGDG in b–e.

Fig. 3. The hydrophilic arm containing NdhV.

a The location of subunits in the hydrophilic domain. The nine NDH subunits in the hydrophilic domain are colored differently and labeled with the abbreviations of one letter (for example, letter V indicates subunit NdhV). The 4Fe–4S clusters are shown as spheres. b Overall structure of NdhV. c The binding position of NdhV. NdhV is shown in cartoon, other subunits are shown in surface mode. NdhI, NdhS, and NdhN are colored differently and labeled, whereas other subunits are shown in white. d The C-terminal helix of NdhV interacts with the hairpin loop of NdhI.

Fig. 4. The Fd binding pocket.

a The Fd binding pocket is mainly formed by NdhI and NdhH. Fd is shown in cartoon, the NDH subunits are shown in surface mode. Subunits close to Fd are colored differently and other subunits are shown in white. b The electrostatic surface representation of Fd binding pocket. Red represents negative charge, and blue represents positive charge. Fd is shown as yellow cartoon. The positively-charged regions surrounding Fd are highlighted with black dashed circle and labeled. c The interactions between Fd and NdhI/NdhH subunits. The Fe–S clusters and the residues involved in the interactions between Fd and NdhI/NdhH are depicted as sticks and labeled. The hydrogen bonds are shown as black dashed lines. The distance between Fe–S clusters of NdhI and Fd (red dashed line) is indicated. d Fd and NdhV almost symmetrically bind the hairpin loop of NdhI from the opposite sides. The zoom-in view of the region circled by the black dashed line is shown in (e). e Superposition of NDH-Fd and NDH-PQ structures showing the different conformation of the hairpin loop in NdhI subunit. Two positively-charged Lys residues from the hairpin loop of NdhI are hydrogen bonded with residues from Fd. Residues involved in the interactions are shown as sticks and labeled. f The loop regions in NdhN and NdhS (pointed with black arrows) interact with NdhI and/or NdhH and help to stabilize the Fd binding pocket.

Fig. 5. The PQ chamber.

a The cofactors involved in the electron transport chain. The PQ molecule is shown as sticks and the Fe–S clusters are shown as spheres, and the distances between cofactors are indicated. The subunits around PQ chamber are shown in surface with the same color code as that in Fig. 1. NdhH subunit is omitted for clarity. b The PQ chamber is shaped by NdhH, NdhK, and NdhA, which are further stabilized by NdhI, NdhL, and NdhC. The structural elements involved in forming the PQ chamber are indicated. The four-helix bundle of NdhH is highlighted with black dashed circle. The PQ molecule is shown in ball-and-stick mode and colored orange. c The interactions around PQ molecules. The N2 cluster, PQ, and key residues are shown in sticks and labeled. Comparison of the PQ chamber in NDH-PQ structure (d) and the Q chamber in Tt complex I structure (PDB ID: 4HEA) (e) in the same view. In NDH-PQ structure, the β1-2 loop containing M22 of NdhH intrudes into the PQ chamber and blocks the path from PQ to Y72 of NdhH and N2 cluster of NdhK, resulting in a smaller occluded PQ chamber (d), while the corresponding loop in Tt complex I structure (colored green) adopts a different conformation and results in a larger opened Q chamber (e). The NDH-PQ structure (colored orange) is superposed to Tt complex I structure (colored green) in e, to show the positions of β1-2 loop and Y72 of NdhH as well as the PQ molecule in NDH-PQ structure. The white arrow indicates that a quinone molecule at the PQ position is able to move toward the N2 cluster in the larger PQ pocket as shown in the Tt complex I structure. f Comparison of the PQ/Q chambers in NDH-PQ, Tt complex I (PDB ID: 4HEA), and Yl complex I (PDB ID: 4WZ7) structures. The β1-2 loop of NdhH in NDH-PQ structure and the corresponding loops in Tt and Yl complex I structures are highlighted in different colors and labeled.

Fig. 6. Comparison of three NDH-1L structures under different pH conditions.

a Comparison of NDH-PQ structure (NDH_pH7) with previously reported two NDH-1L structures NDH_pH6 and NDH_pH8 (PDB codes: 6NBY and 6HUM) superposed on NdhA subunit. The subunits with structural differences are depicted as cartoons and labeled. The regions with evident differences among the three structures are circled with black, blue, and red boxes. b Comparison of the NdhI and NdhH subunits. The N-helix of NdhI and a helix of NdhH are highlighted as cartoons. The subunits similar in three NDH-1L structures are shown as surface mode (For clarity, only NDH_pH7 structure is shown). c Comparison of NdhC subunits viewed from the stromal side. The TMH1-2 loop was not modeled in NDH_pH6 structure. d Comparison of NdhG subunits viewed from the stromal side. Its C-terminal tail in NDH_pH6 structure is almost completely modeled and interacts with the stromal region of NdhB. Color codes are the same as that in Fig. 1.