Photosynthetic Bacterium Rhodobacter Capsulatus Research Articles

Translation Enhancement by a Short Nucleotide Insertion at 5′UTR: Application to an In Vitro Cell-Free System and a Photosynthetic Bacterium

We previously showed that insertion of Dictyostelium gene sequences, such as mlcR, upstream of the Shine–Dalgarno sequence, positively impacts downstream gene expression in Escherichia coli. However, the mechanism by which protein production is facilitated and its applicability to other bacteria remains unknown. In this study, a translation-enhancing effect, associated with this system, on the mRNA amount and property as well as the versatility of the method has been demonstrated. The insertion of mlcR-terminal 25 bp (mlcR25) stabilized the mRNAs and led to increased mRNA levels in E. coli. In the in vitro translation system, a four-fold enhancement was observed when DNA was used as the template, and a three-fold enhancement was observed when mRNA was used as the template. This suggests that mlcR25 has an effect on the facilitation of the interaction between mRNA and ribosome. Furthermore, when this enhancement system was adapted to the photosynthetic bacterium Rhodobacter capsulatus, a more than six-fold increase in translation was observed. Thus, we propose that enhanced translation by mlcR25 is mediated by mechanisms that help the translation machinery to work efficiently, and the system can be applied to bacteria other than E. coli.

Structural Analyses of CrtJ and Its B12-Binding Co-Regulators SAerR and LAerR from the Purple Photosynthetic Bacterium Rhodobacter capsulatus.

Among purple photosynthetic bacteria, the transcription factor CrtJ is a major regulator of photosystem gene expression. Depending on growing conditions, CrtJ can function as an aerobic repressor or an anaerobic activator of photosystem genes. Recently, CrtJ’s activity was shown to be modulated by two size variants of a B12 binding co-regulator called SAerR and LAerR in Rhodobacter capsulatus. The short form, SAerR, promotes CrtJ repression, while the longer variant, LAerR, converts CrtJ into an activator. In this study, we solved the crystal structure of R. capsulatus SAerR at a 2.25 Å resolution. Hydroxycobalamin bound to SAerR is sandwiched between a 4-helix bundle cap, and a Rossman fold. This structure is similar to a AerR-like domain present in CarH from Thermus termophilus, which is a combined photoreceptor/transcription regulator. We also utilized AlphaFold software to predict structures for the LAerR, CrtJ, SAerR-CrtJ and LAerR-CrtJ co-complexes. These structures provide insights into the role of B12 and an LAerR N-terminal extension in regulating the activity of CrtJ.

Persulfide-Responsive Transcription Factor SqrR Regulates Gene Transfer and Biofilm Formation via the Metabolic Modulation of Cyclic di-GMP in Rhodobacter capsulatus.

Bacterial phage-like particles (gene transfer agents—GTAs) are widely employed as a crucial genetic vector in horizontal gene transfer. GTA-mediated gene transfer is induced in response to various stresses; however, regulatory mechanisms are poorly understood. We found that the persulfide-responsive transcription factor SqrR may repress the expression of several GTA-related genes in the photosynthetic bacterium Rhodobacter capsulatus. Here, we show that the sqrR deletion mutant (ΔsqrR) produces higher amounts of intra- and extracellular GTA and gene transfer activity than the wild type (WT). The transcript levels of GTA-related genes are also increased in ΔsqrR. In spite of the presumption that GTA-related genes are regulated in response to sulfide by SqrR, treatment with sulfide did not alter the transcript levels of these genes in the WT strain. Surprisingly, hydrogen peroxide increased the transcript levels of GTA-related genes in the WT, and this alteration was abolished in the ΔsqrR strain. Moreover, the absence of SqrR changed the intracellular cyclic dimeric GMP (c-di-GMP) levels, and the amount of c-di-GMP was correlated with GTA activity and biofilm formation. These results suggest that SqrR is related to the repression of GTA production and the activation of biofilm formation via control of the intracellular c-di-GMP levels.

Reliable determination of the growth and hydrogen production parameters of the photosynthetic bacterium Rhodobacter capsulatus in fed batch culture using a combination of the Gompertz function and the Luedeking-Piret model

In this study, experimental results of hydrogen producing process based on anaerobic photosynthesis using the purple non-sulfur bacterium Rhodobacter capsulatus are scrutinized. The bacterial culture was carried out in a photo-bioreactor operated in a quasi-continuous mode, using lactate as a carbon source. The method is based on the continuous stirred tank reactors (CSTR) technique to access kinetic parameters. The dynamic evolution of hydrogen production as a function of time was accurately simulated using Luedeking-Piret model and the growth of R. capsulatus was computed using Gompertz model. The combination of both models was successfully applied to determine the relevant parameters (λ, μmax, α and β) for two R. capsulatus strains studied: the wild-type strain B10 and the H2 over-producing mutant IR3. The mathematical description indicates that the photofermentation is more promising than dark fermentation for the conversion of organic substrates into biogas.

Structural and electrochemical characterization of lawsone-dependent production of tellurium-metal nanoprecipitates by photosynthetic cells of Rhodobacter capsulatus

Cells of the facultative photosynthetic bacterium Rhodobacter capsulatus exploit the simultaneous presence in the cultural medium of the toxic oxyanion tellurite (TeO32−) and the redox mediator lawsone (2-hydroxy-1,4-naphthoquinone) by reducing tellurite to metal Te0 nanoprecipitates (TeNPs) outside the cells. Here we have studied the mechanism by which lawsone interacts with metabolically active cells and analysed both structure and composition of the TeNPs collected from the growth medium of phototrophycally grown R. capsulatus. High Resolution Transmission Electron Microscopy (HR-TEM) images and Energy-Dispersive X-ray (EDX) microanalysis of TeNPs showed a central core of polycrystalline tellurium interspersed in an organic matrix with a predominant protein-based composition. The main proteins from Te0 nanostructures were identified by Liquid Chromatography tandem-Mass Spectrometry and were all correlated with the cell outer membrane composition. The interaction of reduced lawsone with tellurite and with the bacterial cells was probed by Cyclic Voltammetry and Scanning ElectroChemical Microscopy (SECM). We concluded that lawsone is required for the reduction of tellurite to metal Te0 in a reaction mechanism dependent on reducing equivalents deriving from the cell photosynthetic metabolism. SECM experiments demonstrate that lawsone, by diffusing inside the bacterial cells, is effectively available at the membrane site of the photosynthetic electron transport chain.

The photosynthetic bacteria Rhodobacter capsulatus and Synechocystis sp. PCC 6803 as new hosts for cyclic plant triterpene biosynthesis.

Cyclic triterpenes constitute one of the most diverse groups of plant natural products. Besides the intriguing biochemistry of their biosynthetic pathways, plant triterpenes exhibit versatile bioactivities, including antimicrobial effects against plant and human pathogens. While prokaryotes have been extensively used for the heterologous production of other classes of terpenes, the synthesis of cyclic triterpenes, which inherently includes the two-step catalytic formation of the universal linear precursor 2,3-oxidosqualene, is still a major challenge. We thus explored the suitability of the metabolically versatile photosynthetic α-proteobacterium Rhodobacter capsulatus SB1003 and cyanobacterium Synechocystis sp. PCC 6803 as alternative hosts for biosynthesis of cyclic plant triterpenes. Therefore, 2,3-oxidosqualene production was implemented and subsequently combined with different cyclization reactions catalyzed by the representative oxidosqualene cyclases CAS1 (cycloartenol synthase), LUP1 (lupeol synthase), THAS1 (thalianol synthase) and MRN1 (marneral synthase) derived from model plant Arabidopsis thaliana. While successful accumulation of 2,3-oxidosqualene could be detected by LC-MS analysis in both hosts, cyclase expression resulted in differential production profiles. CAS1 catalyzed conversion to only cycloartenol, but expression of LUP1 yielded lupeol and a triterpenoid matching an oxidation product of lupeol, in both hosts. In contrast, THAS1 expression did not lead to cyclic product formation in either host, whereas MRN1-dependent production of marnerol and hydroxymarnerol was observed in Synechocystis but not in R. capsulatus. Our findings thus indicate that 2,3-oxidosqualene cyclization in heterologous phototrophic bacteria is basically feasible but efficient conversion depends on both the respective cyclase enzyme and individual host properties. Therefore, photosynthetic α-proteo- and cyanobacteria are promising alternative candidates for providing new bacterial access to the broad class of triterpenes for biotechnological applications.

Rapid C8-vinyl reduction of divinyl-chlorophyllide a by BciA from Rhodobacter capsulatus

Abstract Divinyl-chlorophyllide a (DV-Chlide a ) is a universal precursor for chlorophyll or bacteriochlorophyll biosynthesis in all photosynthetic organisms. Previous mutational analyses revealed that BciA works for reduction of the C8-vinyl group of DV-Chlide a. Chlorophyllide oxidoreductase (COR) reduces the C7=C8 double bond of Chlide a bearing the C8-ethyl group, but also potentially catalyzes the reduction of the C8-vinyl group of DV-Chlide a. In this study, we prepared a recombinant BciA protein from the purple photosynthetic bacterium Rhodobacter capsulatus and analyzed its C8-vinyl reduction activity towards DV-Chlide a . BciA formed a functional oligomeric complex consisting of at least three rigid dimers. BciA required NADPH as an electron donor for its C8-vinyl reduction activity. The enzymatic activity of BciA towards the substrate DV-Chlide a was much higher than that of COR towards Chlide a . Phylogenetic distribution and the enzymatic parameters of BciA and COR suggest that BciA is a more recently acquired auxiliary C8-vinyl reductase, attained to meet the high demand for bacteriochlorophylls used to produce large amounts of light-harvesting complexes.

Characterization of redox-active cysteine residues of persulfide-responsive transcriptional repressor SqrR

ABSTRACTWe recently identified the novel persulfide sensor SqrR that functions as a master regulator of sulfide-dependent gene expression in the purple photosynthetic bacterium Rhodobacter capsulatus. SqrR binds to the promoter regions of target genes to repress their expression in the absence of sulfide, and the repressor activity is negated by sulfide treatment. SqrR has 3 cysteine residues, 2 of which are conserved in SqrR homologs from other bacteria: Cys41 and Cys107. SqrR forms an intramolecular tetrasulfide bond between Cys41 and Cys107 when exposed to persulfide, which results in loss of the DNA-binding activity in vitro. Here, we address the mechanism through which these cysteine residues are modified by persulfides. We show that the predicted pKa value of Cys107, as revealed by a putative SqrR structural model, is lower than that of Cys41. Furthermore, C41S SqrR in which Cys41 was changed to serine forms an intermolecular disulfide-bond between Cys107 of 2 SqrRs, suggesting high nucleophilic reactivity of Cy107. These data suggest that Cys107 and Cys41 function as attacking Cys and resolving Cys, respectively; this occurs during tetrasulfide-bond formation of WT SqrR, when it is exposed to persulfide.

Sulfide-responsive transcriptional repressor SqrR functions as a master regulator of sulfide-dependent photosynthesis

Sulfide was used as an electron donor early in the evolution of photosynthesis, with many extant photosynthetic bacteria still capable of using sulfur compounds such as hydrogen sulfide (H2S) as a photosynthetic electron donor. Although enzymes involved in H2S oxidation have been characterized, mechanisms of regulation of sulfide-dependent photosynthesis have not been elucidated. In this study, we have identified a sulfide-responsive transcriptional repressor, SqrR, that functions as a master regulator of sulfide-dependent gene expression in the purple photosynthetic bacterium Rhodobacter capsulatus SqrR has three cysteine residues, two of which, C41 and C107, are conserved in SqrR homologs from other bacteria. Analysis with liquid chromatography coupled with an electrospray-interface tandem-mass spectrometer reveals that SqrR forms an intramolecular tetrasulfide bond between C41 and C107 when incubated with the sulfur donor glutathione persulfide. SqrR is oxidized in sulfide-stressed cells, and tetrasulfide-cross-linked SqrR binds more weakly to a target promoter relative to unmodified SqrR. C41S and C107S R. capsulatus SqrRs lack the ability to respond to sulfide, and constitutively repress target gene expression in cells. These results establish that SqrR is a sensor of H2S-derived reactive sulfur species that maintain sulfide homeostasis in this photosynthetic bacterium and reveal the mechanism of sulfide-dependent transcriptional derepression of genes involved in sulfide metabolism.

Pathways of Transmembrane Electron Transfer in Cytochrome bc Complexes: Dielectric Heterogeneity and Interheme Coulombic Interactions.

The intramembrane cytochrome bc1 complex of the photosynthetic bacterium Rhodobacter capsulatus and the cytochrome b6f complex, which functions in oxygenic photosynthesis, utilize two pairs of b-hemes in a symmetric dimer to accomplish proton-coupled electron transfer. The transmembrane electron transfer pathway in each complex was identified through the novel use of heme Soret band excitonic circular dichroism (CD) spectra, for which the responsible heme-heme interactions were determined from crystal structures. Kinetics of heme reduction and CD amplitude change were measured simultaneously. For bc1, in which the redox potentials of the transmembrane heme pair are separated by 160 mV, heme reduction occurs preferentially to the higher-potential intermonomer heme pair on the electronegative (n) side of the complex. This contrasts with the b6f complex, where the redox potential difference between transmembrane intramonomer p- and n-side hemes is substantially smaller and the n-p pair is preferentially reduced. Limits on the dielectric constant between intramonomer hemes were calculated from the interheme distance and the redox potential difference, ΔEm. The difference in preferred reduction pathway is a consequence of the larger ΔEm between n- and p-side hemes in bc1, which favors the reduction of n-side hemes and cannot be offset by decreased repulsive Coulombic interactions between intramonomer hemes.

Reprint of “Extracellular production of tellurium nanoparticles by the photosynthetic bacterium Rhodobacter capsulatus”

The toxic oxyanion tellurite (TeO32−) is acquired by cells of Rhodobacter capsulatus grown anaerobically in the light, via acetate permease ActP2 and then reduced to Te0 in the cytoplasm as needle-like black precipitates. Interestingly, photosynthetic cultures of R. capsulatus can also generate Te0 nanoprecipitates (TeNPs) outside the cells upon addition of the redox mediator lawsone (2-hydroxy-1,4-naphtoquinone). TeNPs generation kinetics were monitored to define the optimal conditions to produce TeNPs as a function of various carbon sources and lawsone concentration. We report that growing cultures over a 10 days period with daily additions of 1mM tellurite led to the accumulation in the growth medium of TeNPs with dimensions from 200 up to 600–700nm in length as determined by atomic force microscopy (AFM). This result suggests that nucleation of TeNPs takes place over the entire cell growth period although the addition of new tellurium Te0 to pre-formed TeNPs is the main strategy used by R. capsulatus to generate TeNPs outside the cells. Finally, X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared (FT-IR) analysis of TeNPs indicate they are coated with an organic material which keeps the particles in solution in aqueous solvents.

Development of a Rhodobacter capsulatus self-reporting model system for optimizing light-dependent, [FeFe]-hydrogenase-driven H2 production.

The photosynthetic bacterium Rhodobacter capsulatus normally photoproduces H2 as a by-product of its nitrogenase-catalyzed nitrogen-fixing activity. Such H2 production, however, is expensive from a metabolic perspective, requiring nearly four times as many photons as the equivalent algal hydrogenase-based system (Ghirardi et al., 2009 Photobiological hydrogen-producing systems. Chem Soc Rev 38(1):52-61). Here, we report the insertion of a Clostridium acetobutylicum [FeFe]-hydrogenase and its three attendant hydrogenase assembly proteins into an R. capsulatus strain lacking its native uptake hydrogenase. Further, this strain is modified to fluoresce upon sensing H2 . The resulting strain photoproduces H2 and self-reports its own H2 production through fluorescence. This model system represents a unique method of developing hydrogenase-based H2 production in R. capsulatus, may serve as a powerful system for in vivo directed evolution of hydrogenases and hydrogenase-associated genes, and provides a means of screening for increased metabolic production of H2 . Biotechnol. Bioeng. 2017;114: 291-297. © 2016 Wiley Periodicals, Inc.

A kinetic study of hydrogen production by a Calvin-Benson-Bassham cycle mutant, PRK (phosphoribulose kinase), of the photosynthetic bacterium Rhodobacter capsulatus

In the purple non-sulfur photosynthetic bacteria, the Calvin-Benson-Bassham (CBB) cycle plays a role in maintaining intracellular redox balance under photoheterotrophic growth conditions, potentially consuming reductant and chemical input (ATP) that could be used for H2 production by nitrogenase. Previous studies have shown Rhodobacter capsulatus JP91 to be a remarkable H2 producer and in the present study a PRK, phosphoribulose kinase, knockout mutant of this strain was created in an attempt to further improve H2 yield. Analysis of growth with different carbon sources showed that this mutant could only grow in acetate medium without, however, producing any H2. A spontaneous mutant was recovered which had gained the ability to grow on glucose and produce H2 with a yield that was 20–40% greater than the wild type, JP91 strain. Analysis shows that growth and hydrogen production are strongly linked with electrons from glucose mostly directed to H2 production and biomass formation.

Extracellular production of tellurium nanoparticles by the photosynthetic bacterium Rhodobacter capsulatus

The toxic oxyanion tellurite (TeO32−) is acquired by cells of Rhodobacter capsulatus grown anaerobically in the light, via acetate permease ActP2 and then reduced to Te0 in the cytoplasm as needle-like black precipitates. Interestingly, photosynthetic cultures of R. capsulatus can also generate Te0 nanoprecipitates (TeNPs) outside the cells upon addition of the redox mediator lawsone (2-hydroxy-1,4-naphtoquinone). TeNPs generation kinetics were monitored to define the optimal conditions to produce TeNPs as a function of various carbon sources and lawsone concentration. We report that growing cultures over a 10 days period with daily additions of 1mM tellurite led to the accumulation in the growth medium of TeNPs with dimensions from 200 up to 600–700nm in length as determined by atomic force microscopy (AFM). This result suggests that nucleation of TeNPs takes place over the entire cell growth period although the addition of new tellurium Te0 to pre-formed TeNPs is the main strategy used by R. capsulatus to generate TeNPs outside the cells. Finally, X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared (FT-IR) analysis of TeNPs indicate they are coated with an organic material which keeps the particles in solution in aqueous solvents.

Hydrogen Production by a Photosynthetic Bacterium: Some Analytical Solutions

AbstractThe mathematical model for hydrogen production by the photosynthetic bacterium Rhodobacter capsulatus in a batch photobioreactor is discussed. An analytical method – the homotopy perturbation method – is presented to solve the nonlinear differential equations that describe the biomass formation, substrate utilization, and hydrogen production with respect to time. Approximate analytical expressions for the concentrations of biomass, substrate and hydrogen are derived for various values of the relevant parameters. The analytical results are compared with experimental data. In addition, the sensitivity of the kinetic parameters is also analyzed. The time required to achieve the maximum growth of biomass and complete degradation of the substrate is reported.

Clarifying the Roles of Cardiolipin

Cardiolipins (CL) are uniquely structured double lipids that are universally found in membranes which couple electron transport and phosphorylation. The proposed roles of CL in these membranes include mainly two aspects: the effects on the structure and dynamics of the membranes' lipid component and the interplay with membrane-associated proteins. In previous work, we have studied both of these aspects by utilizing atomistic molecular dynamics simulations, starting with the general effects of CL on membrane properties and the interplay with ions [1], further moving on to the interactions with the membrane-embedded respiratory complex cyt bc1 [2]. The latter model system that is of greater interest comprises the entire cyt bc1 dimer of the purple photosynthetic bacterium Rhodobacter capsulatus embedded in a lipid bilayer, whose lipid composition mimics that of the inner mitochondrial membrane. Intriguingly CLs were observed to diffuse spontaneously to the dimer interface and to the immediate vicinity of the catalytic Qi-sites [2]. This observation is in agreement with experimental data, as CLs are indeed located close to the Qi-sites in several X-ray crystal structures of the complex. Importantly, our observations support the proposed role of CL in delivering protons for the non-reduced substrate forms in the active site.In ongoing work that we discuss here we focus more specifically on the roles of individual components of the proposed proton uptake pathway (CL, water, and individual protein residues) and on the atom-level reaction mechanism in the binding pocket. To this end, further MD simulations, QM calculations, and cite-directed mutagenesis experiments were employed. We also discuss whether there is a plausible pathway for substrate movement between the active sites through the lipid-filled insides of the complex, and the role of oxidative stress in cyt bc1 behavior.

Reduction of chalcogen oxyanions and generation of nanoprecipitates by the photosynthetic bacterium Rhodobacter capsulatus

The facultative photosynthetic bacterium Rhodobacter capsulatus is characterized in its interaction with the toxic oxyanions tellurite (TeIV) and selenite (SeIV) by a highly variable level of resistance that is dependent on the growth mode making this bacterium an ideal organism for the study of the microbial interaction with chalcogens. As we have reported in the past, while the oxyanion tellurite is taken up by R. capsulatus cells via acetate permease and it is reduced to Te0 in the cytoplasm in the form of splinter-like black intracellular deposits no clear mechanism was described for Se0 precipitation. Here, we present the first report on the biotransformation of tellurium and selenium oxyanions into extracellular Te0 and Se0nanoprecipitates (NPs) by anaerobic photosynthetically growing cultures of R. capsulatus as a function of exogenously added redox-mediator lawsone, i.e. 2-hydroxy-1,4-naphthoquinone. The NPs formation was dependent on the carbon source used for the bacterial growth and the rate of chalcogen reduction was constant at different lawsone concentrations, in line with a catalytic role for the redox mediator. X-ray diffraction (XRD) analysis demonstrated the Te0 and Se0 nature of the nanoparticles.

Atomistic simulations indicate cardiolipin to have an integral role in the structure of the cytochrome bc1 complex

The reaction mechanism of the cytochrome (cyt) bc1 complex relies on proton and electron transfer to/from the substrate quinone/quinol, which in turn generate a proton gradient across the mitochondrial membrane. Cardiolipin (CL) have been suggested to play an important role in cyt bc1 function by both ensuring the structural integrity of the protein complex and also by taking part in the proton uptake. Yet, the atom-scale understanding of these highly charged four-tail lipids in the cyt bc1 function has remained quite unclear. We consider this issue through atomistic molecular dynamics simulations that are applied to the entire cyt bc1 dimer of the purple photosynthetic bacterium Rhodobacter capsulatus embedded in a lipid bilayer. We find CLs to spontaneously diffuse to the dimer interface to the immediate vicinity of the higher potential heme b groups of the complex's catalytic Qi-sites. This observation is in full agreement with crystallographic studies of the complex, and supports the view that CLs are key players in the proton uptake. The simulation results also allow us to present a refined picture for the dimer arrangement in the cyt bc1 complex, the novelty of our work being the description of the role of the surrounding lipid environment: in addition to the specific CL–protein interactions, we observe the protein domains on the positive side of the membrane to settle against the lipids. Altogether, the simulations discussed in this article provide novel views into the dynamics of cyt bc1 with lipids, complementing previous experimental findings.

Bacillus megaterium has both a functional BluB protein required for DMB synthesis and a related flavoprotein that forms a stable radical species.

Despite the extensive study of the biosynthesis of the complex molecule B12 (cobalamin), the mechanism by which the lower ligand 5,6-dimethylbenzimidazole (DMB) is formed has remained something of a mystery. However, recent work has identified and characterized a DMB-synthase (BluB) responsible for the oxygen-dependent, single enzyme conversion of FMN to DMB. In this work, we have identified BluB homologs from the aerobic purple, nonsulfur, photosynthetic bacterium Rhodobacter capsulatus and the aerobic soil bacterium Bacillus megaterium and have demonstrated DMB synthesis by the use of a novel complementation assay in which a B12 deficient strain, substituted with the precursor cobinamide is recovered either by the addition of DMB or by the recombinant expression of a bluB gene. The DMB-synthetic activity of the purified recombinant BluB enzymes was further confirmed in vitro by providing the enzyme with FMNH2 and oxygen and observing the formation of DMB by HPLC. The formation of a 4a-peroxyflavin intermediate, the first step in the oxygen dependent mechanism of DMB biosynthesis, is reported here and is the first intermediate in the enzyme catalysed reaction to be demonstrated experimentally to date. The identification and characterization of an FMN-binding protein found on the cobI operon of B. megaterium, CbiY, is also detailed, revealing an FMN-containing enzyme which is able to stabilize a blue flavin semiquinone upon reduction with a 1-electron donor.

Atomistic Simulations Indicate Cardiolipin to have an Integral Role in the Structure of the Cytochrome bc1 Complex

The reaction mechanism of the cytochrome bc1 (cyt bc1) complex relies on proton and electron transfers to/from substrate quinone/quinol, which in turn generates a proton gradient across the mitochondrial membrane used in the ATP synthesis. Cardiolipins (CLs) have been suggested to both ensure the structural integrity of the complex and take part in the proton uptake. We study the issue by simulating the entire cyt bc1 dimer of purple photosynthetic bacterium Rhodobacter capsulatus embedded into a lipid bilayer using all-atom molecular dynamics. In the 200 ns simulations CLs position themselves in the dimer interface and around the protein close to the higher potential heme groups of the enzyme complex's catalytic Qi-sites, in line with the positioning seen in the crystal structures. The arrangement of CLs close to the Qi-sites supports the proposed view of these highly charged phospholipids as key players in the proton uptake. Based on the simulations we also present a refined dimer arrangement for the cyt bc1 complex that encompasses the full effect of the surrounding lipid environment. The periplasmic domains of the complex spread against the membrane; moreover, this conformational shift was more pronounced when Qo-site lacked a bound substrate. As a whole, the applied modeling approach provides novel insights into the lipid-cyt bc1 dynamics, complementing the previous experimental findings.