Cyanobacterial NDH-1 Complexes Research Articles

Cyanobacterial NDH-1 Complexes.

Light reaction of photosynthesis is efficiently driven by protein complexes arranged in an orderly in the thylakoid membrane. As the 5th complex, NAD(P)H dehydrogenase complex (NDH-1) is involved in cyclic electron flow around photosystem I to protect plants against environmental stresses for efficient photosynthesis. In addition, two kinds of NDH-1 complexes participate in CO2 uptake for CO2 concentration in cyanobacteria. In recent years, great progress has been made in the understanding of the assembly and the structure of NDH-1. However, the regulatory mechanism of NDH-1 in photosynthesis remains largely unknown. Therefore, understanding the regulatory mechanism of NDH-1 is of great significance to reveal the mechanism of efficient photosynthesis. In this mini-review, the author introduces current progress in the research of cyanobacterial NDH-1. Finally, the author summarizes the possible regulatory mechanism of cyanobacterial NDH-1 in photosynthesis and discusses the research prospect.

Electron flow through NDH-1 complexes is the major driver of cyclic electron flow-dependent proton pumping in cyanobacteria

Cyclic electron flow (CEF) around photosystem I is vital to balancing the photosynthetic energy budget of cyanobacteria and other photosynthetic organisms. The coupling of CEF to proton pumping has long been hypothesized to occur, providing proton motive force (PMF) for the synthesis of ATP with no net cost to [NADPH]. This is thought to occur largely through the activity of NDH-1 complexes, of which cyanobacteria have four with different activities. While a much work has been done to understand the steady-state PMF in both the light and dark, and fluorescent probes have been developed to observe these fluxes in vivo, little has been done to understand the kinetics of these fluxes, particularly with regard to NDH-1 complexes. To monitor the kinetics of proton pumping in Synechocystis sp. PCC 6803, the pH sensitive dye Acridine Orange was used alongside a suite of inhibitors in order to observe light-dependent proton pumping. The assay was demonstrated to measure photosynthetically driven proton pumping and used to measure the rates of proton pumping unimpeded by dark ΔpH. Here, the cyanobacterial NDH-1 complexes are shown to pump a sizable portion of proton flux when CEF-driven and LEF-driven proton pumping rates are observed and compared in mutants lacking some or all NDH-1 complexes. It is also demonstrated that PSII and LEF are responsible for the bulk of light induced proton pumping, though CEF and NDH-1 are capable of generating ~40% of the proton pumping rate when LEF is inactivated.

Molecular dynamics and structural models of the cyanobacterial NDH-1 complex

NDH-1 is a gigantic redox-driven proton pump linked with respiration and cyclic electron flow in cyanobacterial cells. Based on experimentally resolved X-ray and cryo-EM structures of the respiratory complex I, we derive here molecular models of two isoforms of the cyanobacterial NDH-1 complex involved in redox-driven proton pumping (NDH-1L) and CO2-fixation (NDH-1MS). Our models show distinct structural and dynamic similarities to the core architecture of the bacterial and mammalian respiratory complex I. We identify putative plastoquinone-binding sites that are coupled by an electrostatic wire to the proton pumping elements in the membrane domain of the enzyme. Molecular simulations suggest that the NDH-1L isoform undergoes large-scale hydration changes that support proton-pumping within antiporter-like subunits, whereas the terminal subunit of the NDH-1MS isoform lacks such structural motifs. Our work provides a putative molecular blueprint for the complex I-analogue in the photosynthetic energy transduction machinery and demonstrates that general mechanistic features of the long-range proton-pumping machinery are evolutionary conserved in the complex I-superfamily.

NdhV Is a Subunit of NADPH Dehydrogenase Essential for Cyclic Electron Transport in Synechocystis sp. Strain PCC 6803.

Two mutants sensitive to heat stress for growth and impaired in NADPH dehydrogenase (NDH-1)-dependent cyclic electron transport around photosystem I (NDH-CET) were isolated from the cyanobacterium Synechocystis sp. strain PCC 6803 transformed with a transposon-bearing library. Both mutants had a tag in the same sll0272 gene, encoding a protein highly homologous to NdhV identified in Arabidopsis (Arabidopsis thaliana). Deletion of the sll0272 gene (ndhV) did not influence the assembly of NDH-1 complexes and the activities of CO2 uptake and respiration but reduced the activity of NDH-CET. NdhV interacted with NdhS, a ferredoxin-binding subunit of cyanobacterial NDH-1 complex. Deletion of NdhS completely abolished NdhV, but deletion of NdhV had no effect on the amount of NdhS. Reduction of NDH-CET activity was more significant in ΔndhS than in ΔndhV. We therefore propose that NdhV cooperates with NdhS to accept electrons from reduced ferredoxin.

Solution structure of the NDH-1 complex subunit CupS from Thermosynechococcus elongatus

The cyanobacterial multi-subunit membrane protein complex NDH-1 is both structurally and functionally related to Complex I of eubacteria and mitochondria. In addition to functions in respiration and cyclic electron transfer around photosystem I (PSI), the cyanobacterial NDH-1 complex is involved in a unique mechanism for inorganic carbon concentration. Although the crystal structures of the similar respiratory Complex I from Thermus thermophilus and Escherichia coli are known, atomic structural information is not available for the cyanobacterial NDH-1 complex yet. In particular, the structures of those subunits that are not homologous to Complex I will help to understand their distinct functions. The 15.7kDa protein CupS is a small soluble subunit of the complex variant NDH-1MS, which is thought to play a role in CO2 conversion.Here, we present the NMR structure of CupS from Thermosynechococcus elongatus, which is the very first structure of a specific cyanobacterial NDH-1 complex subunit. CupS shares a structural similarity with members of the Fasciclin protein superfamily. The structural comparison to Fasciclin type proteins based on known NMR structures and protein sequences of human TGFBIp, MPB70 from Mycobacterium bovis, and Fdp from Rhodobacter sphaeroides, together with a virtual docking model of CupS and NdhF3, provide first insight into the specific binding of CupS to the NDH-1MS complex at atomic resolution.

NDH-1L interacts with ferredoxin via the subunit NdhS in Thermosynechococcus elongatus.

The large size complex of cyanobacterial NAD(P)H dehydrogenase (NDH-1) complex (NDH-1L) plays crucial role in a variety of bioenergetic reactions such as respiration and cyclic electron flow around photosystem I. Although the complex has been isolated and identified, its biochemical function still remains to be clarified. Here, we highly purified the NDH-1L complex from the cells of Thermosynechococcus elongatus by Ni(2+) affinity chromatography and size-exclusion chromatography. The purified NDH-1L complex has an apparent total molecular mass of approximately 500kDa. 14 known subunits were identified by mass spectrometry and immunoblotting, including the NdhS subunit containing ferredoxin (Fd)-docking site domain. Surface plasmon resonance measurement demonstrates that the NDH-1L complex could bind to Fd with the binding constant (K D) of 59µM. Yeast two-hybrid system assay further confirmed the interaction of Fd with NdhS and indicated that NdhH is involved in the interaction. Our results provide direct biochemical evidence that the cyanobacterial NDH-1 complex catalyzes the electron transport from reduced Fd to plastoquinone via NdhS and NdhH.

Deletion of an electron donor-binding subunit of the NDH-1 complex, NdhS, results in a heat-sensitive growth phenotype in Synechocystis sp. PCC 6803

In cyanobacteria and higher plants, NdhS is suggested to be an electron donor-binding subunit of NADPH dehydrogenase (NDH-1) complexes and its absence impairs NDH-1-dependent cyclic electron transport around photosystem I (NDH-CET). Despite significant advances in the study of NdhS during recent years, its functional role in resisting heat stress is poorly understood. Here, our results revealed that the absence of NdhS resulted in a serious heat-sensitive growth phenotype in the unicellular cyanobacterium Synechocystis sp. strain PCC 6803. Furthermore, the rapid and significant increase in NDH-CET caused by heat treatment was completely abolished, and the repair of photosystem II under heat stress conditions was greatly impaired when compared to that of other photosynthetic apparatus in the thylakoid membrane. We therefore conclude that NdhS plays an important role in resistance to heat stress, possibly by stabilizing the electron input module of cyanobacterial NDH-1 complexes.

(1)H, (13)C and (15)N chemical shift assignments of the NDH-1 complex subunit CupS.

The cyanobacterial NDH-1 complex is involved in respiratory as well as in cyclic electron transfer around photosystem I. Here, we report both backbone and side chain chemical shift assignments of CupS, a small subunit of the multisubunit membrane protein complex NDH-1 from Thermosynechococcus elongatus. The construct contains 159 amino acids including a Strep-tag and two additional amino acids.

The Gene sml0013 of Synechocystis Species Strain PCC 6803 Encodes for a Novel Subunit of the NAD(P)H Oxidoreductase or Complex I That Is Ubiquitously Distributed among Cyanobacteria

The NAD(P)H oxidoreductase or complex I (NDH1) complex participates in many processes such as respiration, cyclic electron flow, and inorganic carbon concentration in the cyanobacterial cell. Despite immense progress in our understanding of the structure-function relation of the cyanobacterial NDH1 complex, the subunits catalyzing NAD(P)H docking and oxidation are still missing. The gene sml0013 of Synechocystis 6803 encodes for a small protein of unknown function for which homologs exist in all completely known cyanobacterial genomes. The protein exhibits weak similarities to the NDH-dependent flow6 (NDF6) protein, which was reported from Arabidopsis (Arabidopsis thaliana) chloroplasts as a NDH subunit. An sml0013 inactivation mutant of Synechocystis 6803 was generated and characterized. It showed only weak differences regarding growth and pigmentation in various culture conditions; most remarkably, it exhibited a glucose-sensitive phenotype in the light. The genome-wide expression pattern of the Δsml0013::Km mutant was almost identical to the wild type when grown under high CO2 conditions as well as after shifts to low CO2 conditions. However, measurements of the photosystem I redox kinetic in cells of the Δsml0013::Km mutant revealed differences, such as a decreased capability of cyclic electron flow as well as electron flow into respiration in comparison with the wild type. These results suggest that the Sml0013 protein (named NdhP) represents a novel subunit of the cyanobacterial NDH1 complex, mediating its coupling either to the respiratory or the photosynthetic electron flow.

Identification of Novel Ssl0352 Protein (NdhS), Essential for Efficient Operation of Cyclic Electron Transport around Photosystem I, in NADPH:plastoquinone Oxidoreductase (NDH-1) Complexes of Synechocystis sp. PCC 6803

Cyanobacterial NADPH:plastoquinone oxidoreductase, or type I NAD(P)H dehydrogenase, or the NDH-1 complex is involved in plastoquinone reduction and cyclic electron transfer (CET) around photosystem I. CET, in turn, produces extra ATP for cell metabolism particularly under stressful conditions. Despite significant achievements in the study of cyanobacterial NDH-1 complexes during the past few years, the entire subunit composition still remains elusive. To identify missing subunits, we screened a transposon-tagged library of Synechocystis 6803 cells grown under high light. Two NDH-1-mediated CET (NDH-CET)-defective mutants were tagged in the same ssl0352 gene encoding a short unknown protein. To clarify the function of Ssl0352, the ssl0352 deletion mutant and another mutant with Ssl0352 fused to yellow fluorescent protein (YFP) and the His(6) tag were constructed. Immunoblotting, mass spectrometry, and confocal microscopy analyses revealed that the Ssl0352 protein resides in the thylakoid membrane and associates with the NDH-1L and NDH-1M complexes. We conclude that Ssl0352 is a novel subunit of cyanobacterial NDH-1 complexes and designate it NdhS. Deletion of the ssl0352 gene considerably impaired the NDH-CET activity and also retarded cell growth under high light conditions, indicating that NdhS is essential for efficient operation of NDH-CET. However, the assembly of the NDH-1L and NDH-1M complexes and their content in the cells were not affected in the mutant. NdhS contains a Src homology 3-like domain and might be involved in interaction of the NDH-1 complex with an electron donor.

Marine cyanophages: tinkering with the electron transport chain

Marine cyanophages: tinkering with the electron transport chain

NdhP and NdhQ: Two Novel Small Subunits of the Cyanobacterial NDH-1 Complex

The subunit composition of the NAD(P)H dehydrogenase complex of Thermosynechococcus elongatus was analyzed by different types of mass spectrometry. All 15 known subunits (NdhA-NdhO) were identified in the purified NDH-1L complex. Moreover, two additional intact mass tags of 4902.7 and 4710.5 Da could be assigned after reannotation of the T. elongatus genome. NdhP and NdhQ are predicted to contain a single transmembrane helix each, and homologues are apparent in other cyanobacteria. Additionally, ndhP is present in some cyanophages in a cluster of PSI genes and exhibits partial similarity to NDF6, a subunit of the plant NDH-1 complex.

Cyanobacterial NDH-1 complexes: Novel insights and remaining puzzles

Cyanobacterial NDH-1 complexes belong to a family of energy converting NAD(P)H:Quinone oxidoreductases that includes bacterial type-I NADH dehydrogenase and mitochondrial Complex I. Several distinct NDH-1 complexes may coexist in cyanobacterial cells and thus be responsible for a variety of functions including respiration, cyclic electron flow around PSI and CO 2 uptake. The present review is focused on specific features that allow to regard the cyanobacterial NDH-1 complexes, together with NDH complexes from chloroplasts, as a separate sub-class of the Complex I family of enzymes. Here, we summarize our current knowledge about structure of functionally different NDH-1 complexes in cyanobacteria and consider implications for a functional mechanism. This article is part of a Special Issue entitled: Regulation of Electron Transport in Chloroplasts.

Identification, regulation and physiological functions of multiple NADPH dehydrogenase complexes in cyanobacteria

Cyanobacteria possess multiple, functionally distinct NADPH dehydrogenase (NDH-1) complexes. In this mini-review, we describe the cyanobacterial NDH-1 complexes by focusing on their identification, regulatory properties, and multiple functions. The multiple functions can be divided into basic and extending functions, and the basic functions are compared with those in chloroplasts. Many questions related to cyanobacterial NDH-1 complexes remain unanswered and are briefly summarized here.

Cyanobacterial NDH‐1 complexes: multiplicity in function and subunit composition

In cyanobacteria, the NAD(P)H:quinone oxidoreductase (NDH-1) is involved in a variety of functions like respiration, cyclic electron flow around PSI and CO(2) uptake. Several types of NDH-1 complexes, which differ in structure and are responsible for these functions, exist in cyanobacterial membranes. This minireview is based on data obtained by reverse genetics and proteomics studies and focuses on the structural and functional differences of the two types of cyanobacterial NDH-1 complexes: NDH-1L, important for respiration and PSI cyclic electron flow, and NDH-1MS, the low-CO(2) inducible complex participating in CO(2) uptake. The NDH-1 complexes in cyanobacteria share a common NDH-1M 'core' complex and differ in the composition of the distal membrane domain composed of specific NdhD and NdhF proteins, which in complexes involved in CO(2) uptake is further associated with the hydrophilic carbon uptake (CUP) domain. At present, however, very important questions concerning the nature of catalytically active subunits that constitute the electron input device (like NADH dehydrogenase module of the eubacterial 'model' NDH-1 analogs), the substrate specificity and reaction mechanisms of cyanobacterial complexes remain unanswered and are shortly discussed here.

Cyanobacterial NADPH dehydrogenase complexes

Cyanobacteria possess functionally distinct multiple NADPH dehydrogenase (NDH-1) complexes that are essential to CO(2) uptake, photosystem-1 cyclic electron transport and respiration. The unique nature of cyanobacterial NDH-1 complexes is the presence of subunits involved in CO(2) uptake. Other than CO(2) uptake, chloroplastic NDH-1 complex has a similar role as cyanobacterial NDH-1 complexes in photosystem-1 cyclic electron transport and respiration (chlororespiration). In this mini-review we focus on the structure and function of cyanobacterial NDH-1 complexes and their phylogeny. The function of chloroplastic NDH-1 complex and characteristics of plants defective in NDH-1 are also described for comparison.