KaiC family ATPases in the nonheterocystous nitrogen-fixing cyanobacterium Leptolyngbya boryana.

A circadian clock is reconstituted in vitro by incubating three proteins, KaiA, KaiB, and KaiC from the non-nitrogen-fixing cyanobacterium Synechococcus elongatus PCC 7942 in the presence of ATP. Leptolyngbya boryana is a filamentous cyanobacterium that grows diazotrophically under microoxic conditions. Among the aforementioned proteins, KaiC is the main clock oscillator belonging to the RecA ATPase superfamily.

Na + -driven pH regulation by Na+/H+ antiporters promotes photosynthetic efficiency in cyanobacteria.

Photosynthetic organisms have developed mechanisms to regulate light reactions in response to varying light conditions. Photosynthetic electron transport leads to the formation of a ΔpH across the thylakoid membrane (TM), which is crucial for regulating electron transport. However, other pH modulators remain to be identified, particularly in cyanobacteria.

Experimental and Theoretical Mutation of Exciton States on the Smallest Type-I Photosynthetic Reaction Center Complex of a Green Sulfur Bacterium .

The exciton states on the smallest type-I photosynthetic reaction center complex of a green sulfur bacterium (GsbRC) consisting of 26 bacteriochlorophylls (BChl ) and four chlorophylls (Chl ) located on the homodimer of two PscA reaction center polypeptides were investigated. This analysis involved the study of exciton states through a combination of theoretical modeling and the genetic removal of BChl pigments at eight sites. (1) A theoretical model of the pigment assembly exciton state on GsbRC was constructed using Poisson TrESP (P-TrESP) and charge density coupling (CDC) methods based on structural information.

Soluble domains of cytochrome -556 and Rieske iron-sulfur protein from m: Crystal structures and interaction analysis.

In photosynthetic green sulfur bacteria, the electron transfer reaction from menaquinol:cytochrome oxidoreductase to the P840 reaction center (RC) complex occurs directly without any involvement of soluble electron carrier protein(s). X-ray crystallography has determined the three-dimensional structures of the soluble domains of the gene product and Rieske iron-sulfur protein (ISP). The former is a mono-heme cytochrome with an α-absorption peak at 556 nm.

Recent advances in the structural diversity of reaction centers.

Photosynthetic reaction centers (RC) catalyze the conversion of light to chemical energy that supports life on Earth, but they exhibit substantial diversity among different phyla. This is exemplified in a recent structure of the RC from an anoxygenic green sulfur bacterium (GsbRC) which has characteristics that may challenge the canonical view of RC classification. The GsbRC structure is analyzed and compared with other RCs, and the observations reveal important but unstudied research directions that are vital for disentangling RC evolution and diversity.

Mutation of alanine-422 in KaiC leads to a low amplitude of rhythm in the reconstituted cyanobacterial circadian clock.

The cyanobacterial circadian oscillator can be reconstituted by mixing the purified clock proteins KaiA, KaiB, and KaiC with ATP in vitro, leading to a 24-h oscillation of KaiC phosphorylation. The cyanobacterial mutant pr1 carrying valine instead of alanine at position 422 of KaiC (KaiC-A422V) lost the ability to shift the phase of the circadian rhythm. In this study, we analyzed KaiC-A422V to investigate the effect of this single-residue substitution on the in vitro reconstitution of KaiC oscillation.

KaiC3 Displays Temperature- and KaiB-Dependent ATPase Activity and Is Important for Growth in Darkness.

Cyanobacteria form a heterogeneous bacterial group with diverse lifestyles, acclimation strategies, and differences in the presence of circadian clock proteins. In PCC 7942, a unique posttranslational KaiABC oscillator drives circadian rhythms. ATPase activity of KaiC correlates with the period of the clock and mediates temperature compensation.

Soft X-Ray Imaging of Cellular Carbon and Nitrogen Distributions in Heterocystous Cyanobacteria.

Soft x-ray microscopy (SXM) is a minimally invasive technique for single-cell high-resolution imaging as well as the visualization of intracellular distributions of light elements such as carbon, nitrogen, and oxygen. We used SXM to observe photosynthesis and nitrogen fixation in the filamentous cyanobacterium sp. PCC 7120, which can form heterocysts during nitrogen starvation.

Light-Induced Electron Spin-Polarized (ESP) EPR Signal of the P800 Menaquinone Radical Pair State in Oriented Membranes of Heliobacterium modesticaldum: Role/Location of Menaquinone in the Homodimeric Type I Reaction Center.

Function/location of menaquinone (MQ) was studied in the photosynthetic reaction center of Heliobacterium (Hbt.) modesticaldum (hRC), which is one of the most primitive homodimeric type I RCs. The spin-polarized electron paramagnetic resonance signals of light-induced radical pair species, which are made of oxidized electron donor bacteriochlorophyll g (P800) and reduced menaquinone (MQ) or iron-sulfur cluster (F), were measured in the oriented membranes of Hbt.

Phosphorylation at Thr432 induces structural destabilization of the CII ring in the circadian oscillator KaiC.

KaiC is the central oscillator protein in the cyanobacterial circadian clock. KaiC oscillates autonomously between phosphorylated and dephosphorylated states on a 24-h cycle in vitro by mixing with KaiA and KaiB in the presence of ATP. KaiC forms a C -symmetrical hexamer, which is a double ring structure of homologous N-terminal and C-terminal domains termed CI and CII, respectively.

Conversion between two conformational states of KaiC is induced by ATP hydrolysis as a trigger for cyanobacterial circadian oscillation.

The cyanobacterial circadian oscillator can be reconstituted in vitro by mixing three clock proteins, KaiA, KaiB and KaiC, with ATP. KaiC is the only protein with circadian rhythmic activities. In the present study, we tracked the complex formation of the three Kai proteins over time using blue native (BN) polyacrylamide gel electrophoresis (PAGE), in which proteins are charged with the anionic dye Coomassie brilliant blue (CBB).

Orientations of Iron-Sulfur Clusters FA and FB in the Homodimeric Type-I Photosynthetic Reaction Center of Heliobacterium modesticaldum.

Orientations of the FA and FB iron-sulfur (FeS) clusters in a structure-unknown type-I homodimeric heriobacterial reaction center (hRC) were studied in oriented membranes of the thermophilic anaerobic photosynthetic bacterium Heliobacterium modesticaldum by electron paramagnetic resonance (EPR), and compared with those in heterodimeric photosystem I (PS I). The Rieske-type FeS center in the cytochrome b/c complex showed a well-oriented EPR signal. Illumination at 14 K induced an FB(-) signal with g-axes of gz = 2.

Mutation-induced perturbation of the special pair P840 in the homodimeric reaction center in green sulfur bacteria.

Homodimeric photosynthetic reaction centers (RCs) in green sulfur bacteria and heliobacteria are functional homologs of Photosystem (PS) I in oxygenic phototrophs. They show unique features in their electron transfer reactions; however, detailed structural information has not been available so far. We mutated PscA-Leu688 and PscA-Val689 to cysteine residues in the green sulfur bacterium Chlorobaculum tepidum; these residues were predicted to interact with the special pair P840, based on sequence comparison with PS I.

Menaquinone as the Secondary Electron Acceptor in the Type I Homodimeric Photosynthetic Reaction Center of Heliobacterium modesticaldum.

The type I photosynthetic reaction center (RC) of heliobacteria (hRC) is a homodimer containing cofactors almost analogous to those in the plant photosystem I (PS I). However, its three-dimensional structure is not yet clear. PS I uses phylloquinone (PhyQ) as a secondary electron acceptor (A1), while the available evidence has suggested that menaquinone (MQ) in hRC has no function as A1.

Gene expression system in green sulfur bacteria by conjugative plasmid transfer.

Gene transfer and expression systems in green sulfur bacteria were established by bacterial conjugation with Escherichia coli. Conjugative plasmid transfer from E. coli S17-1 to a thermophilic green sulfur bacterium, Chlorobaculum tepidum (formerly Chlorobium tepidum) WT2321, was executed with RSF1010-derivative broad-host-range plasmids, named pDSK5191 and pDSK5192, that confer erythromycin and streptomycin/spectinomycin resistance, respectively.

A heterogeneous tag-attachment to the homodimeric type 1 photosynthetic reaction center core protein in the green sulfur bacterium Chlorobaculum tepidum.

The 6xHis-tag-pscA gene, which was genetically engineered to express N-terminally histidine (His)-tagged PscA, was inserted into a coding region of the recA gene in the green sulfur bacterium Chlorobaculum tepidum (C. tepidum). Although the inactivation of the recA gene strongly suppressed a homologous recombination in C.

Crystal structure of the electron carrier domain of the reaction center cytochrome c(z) subunit from green photosynthetic bacterium Chlorobium tepidum.

In green sulfur photosynthetic bacteria, the cytochrome c(z) (cyt c(z)) subunit in the reaction center complex mediates electron transfer mainly from menaquinol/cytochrome c oxidoreductase to the special pair (P840) of the reaction center. The cyt c(z) subunit consists of an N-terminal transmembrane domain and a C-terminal soluble domain that binds a single heme group. The periplasmic soluble domain has been proposed to be highly mobile and to fluctuate between oxidoreductase and P840 during photosynthetic electron transfer.

C-type cytochromes in the photosynthetic electron transfer pathways in green sulfur bacteria and heliobacteria.

Green sulfur bacteria and heliobacteria are strictly anaerobic phototrophs that have homodimeric type 1 reaction center complexes. Within these complexes, highly reducing substances are produced through an initial charge separation followed by electron transfer reactions driven by light energy absorption. In order to attain efficient energy conversion, it is important for the photooxidized reaction center to be rapidly rereduced.

Sulfur oxidation in mutants of the photosynthetic green sulfur bacterium Chlorobium tepidum devoid of cytochrome c-554 and SoxB.

A mutant devoid of cytochrome c-554 (CT0075) in Chlorobium tepidum (syn. Chlorobaculum tepidum) exhibited a decreased growth rate but normal growth yield when compared to the wild type. From quantitative determinations of sulfur compounds in media, the mutant was found to oxidize thiosulfate more slowly than the wild type but completely to sulfate as the wild type.

Accumulation of chlorophyllous pigments esterified with the geranylgeranyl group and photosynthetic competence in the CT2256-deleted mutant of the green sulfur bacterium Chlorobium tepidum.

Green sulfur bacteria contain chlorophyllous pigments, chlorophyll (Chl) aPD and bacteriochlorophyll (BChl) aP, esterified with Delta2,6-phytadienol and phytol, respectively, which would be produced by reduction of the geranylgeranyl group at the C-17 propionate residue. In the genome of Chlorobium tepidum, two paralogous genes presumably encoding geranylgeranyl reductase, CT1232 and CT2256, are found. The deletion mutants of the CT1232 and CT2256 genes were constructed using an insertional inactivation method in order to clarify the biosynthetic process of the Delta2,6-phytadienyl and phytyl groups in green sulfur bacteria.

Parallel electron donation pathways to cytochrome c(z) in the type I homodimeric photosynthetic reaction center complex of Chlorobium tepidum.

We studied the regulation mechanism of electron donations from menaquinol:cytochrome c oxidoreductase and cytochrome c-554 to the type I homodimeric photosynthetic reaction center complex of the green sulfur bacterium Chlorobium tepidum. We measured flash-induced absorption changes of multiple cytochromes in the membranes prepared from a mutant devoid of cytochrome c-554 or in the reconstituted membranes by exogenously adding cytochrome c-555 purified from Chlorobium limicola. The results indicated that the photo-oxidized cytochrome c(z) bound to the reaction center was rereduced rapidly by cytochrome c-555 as well as by the menaquinol:cytochrome c oxidoreductase and that cytochrome c-555 did not function as a shuttle-like electron carrier between the menaquinol:cytochrome c oxidoreductase and cytochrome c(z).