A photoheterotrophic bacterium from Iceland has adapted its photosynthetic machinery to the long days of polar summer.

During their long evolution, anoxygenic phototrophic bacteria have inhabited a wide variety of natural habitats and developed specific strategies to cope with the challenges of any particular environment. Expression, assembly, and safe operation of the photosynthetic apparatus must be regulated to prevent reactive oxygen species generation under illumination in the presence of oxygen. Here, we report on the photoheterotrophic Sediminicoccus sp. strain KRV36, which was isolated from a cold stream in north-western Iceland, 30 km south of the Arctic Circle. In contrast to most aerobic anoxygenic phototrophs, which stop pigment synthesis when illuminated, strain KRV36 maintained its bacteriochlorophyll synthesis even under continuous light. Its cells also contained between 100 and 180 chromatophores, each accommodating photosynthetic complexes that exhibit an unusually large carotenoid absorption spectrum. The expression of photosynthesis genes in dark-adapted cells was transiently downregulated in the first 2 hours exposed to light but recovered to the initial level within 24 hours. An excess of membrane-bound carotenoids as well as high, constitutive expression of oxidative stress response genes provided the required potential for scavenging reactive oxygen species, safeguarding bacteriochlorophyll synthesis and photosystem assembly. The unique cellular architecture and an unusual gene expression pattern represent a specific adaptation that allows the maintenance of anoxygenic phototrophy under arctic conditions characterized by long summer days with relatively low irradiance.IMPORTANCEThe photoheterotrophic bacterium Sediminicoccus sp. KRV36 was isolated from a cold stream in Iceland. It expresses its photosynthesis genes, synthesizes bacteriochlorophyll, and assembles functional photosynthetic complexes under continuous light in the presence of oxygen. Unraveling the molecular basis of this ability, which is exceptional among aerobic anoxygenic phototrophic species, will help to understand the evolution of bacterial photosynthesis in response to changing environmental conditions. It might also open new possibilities for genetic engineering of biotechnologically relevant phototrophs, with the aim of increasing photosynthetic activity and their tolerance to reactive oxygen species.

Intracytoplasmic-membrane development in alphaproteobacteria involves the homolog of the mitochondrial crista-developing protein Mic60.

Mitochondrial cristae expand the surface area of respiratory membranes and ultimately allow for the evolutionary scaling of respiration with cell volume across eukaryotes. The discovery of Mic60 homologs among alphaproteobacteria, the closest extant relatives of mitochondria, suggested that cristae might have evolved from bacterial intracytoplasmic membranes (ICMs). Here, we investigated the predicted structure and function of alphaproteobacterial Mic60, and a protein encoded by an adjacent gene Orf52, in two distantly related purple alphaproteobacteria, Rhodobacter sphaeroides and Rhodopseudomonas palustris. In addition, we assessed the potential physical interactors of Mic60 and Orf52 in R. sphaeroides. We show that the three α helices of mitochondrial Mic60's mitofilin domain, as well as its adjacent membrane-binding amphipathic helix, are present in alphaproteobacterial Mic60. The disruption of Mic60 and Orf52 caused photoheterotrophic growth defects, which are most severe under low light conditions, and both their disruption and overexpression led to enlarged ICMs in both studied alphaproteobacteria. We also found that alphaproteobacterial Mic60 physically interacts with BamA, the homolog of Sam50, one of the main physical interactors of eukaryotic Mic60. This interaction, responsible for making contact sites at mitochondrial envelopes, has been conserved in modern alphaproteobacteria despite more than a billion years of evolutionary divergence. Our results suggest a role for Mic60 in photosynthetic ICM development and contact site formation at alphaproteobacterial envelopes. Overall, we provide support for the hypothesis that mitochondrial cristae evolved from alphaproteobacterial ICMs and have therefore improved our understanding of the nature of the mitochondrial ancestor.

Characterisation of the photosynthetic complexes from the marine gammaproteobacterium Congregibacter litoralis KT71.

Possibly the most abundant group of anoxygenic phototrophs are marine photoheterotrophic Gammaproteobacteria belonging to the NOR5/OM60 clade. As little is known about their photosynthetic apparatus, the photosynthetic complexes from the marine phototrophic bacterium Congregibacter litoralis KT71 were purified and spectroscopically characterised. The intra-cytoplasmic membranes contain a smaller amount of photosynthetic complexes when compared with anaerobic purple bacteria. Moreover, the intra-cytoplasmic membranes contain only a minimum amount of peripheral LH2 complexes. The complexes are populated by bacteriochlorophyll a, spirilloxanthin and two novel ketocarotenoids, with biophysical and biochemical properties similar to previously characterised complexes from purple bacteria. The organization of the RC-LH1 complex has been further characterised using cryo-electron microscopy. The overall organisation is similar to the complex from the gammaproteobacterium Thermochromatium tepidum, with the type-II reaction centre surrounded by a slightly elliptical LH1 antenna ring composed of 16 αß-subunits with no discernible gap or pore. The RC-LH1 and LH2 apoproteins are phylogenetically related to other halophilic species but LH2 also to some alphaproteobacterial species. It seems that the reduction of light-harvesting apparatus and acquisition of novel ketocarotenoids in Congregibacter litoralis KT71 represent specific adaptations for operating the anoxygenic photosynthesis under aerobic conditions at sea.

Quantum chemical elucidation of a sevenfold symmetric bacterial antenna complex.

The light-harvesting complex 2 (LH2) of purple bacteria is one of the most studied photosynthetic antenna complexes. Its symmetric structure and ring-like bacteriochlorophyll arrangement make it an ideal system for theoreticians and spectroscopists. LH2 complexes from most bacterial species are thought to have eightfold or ninefold symmetry, but recently a sevenfold symmetric LH2 structure from the bacterium Mch. purpuratum was solved by Cryo-Electron microscopy. This LH2 also possesses unique near-infrared absorption and circular dichroism (CD) spectral properties. Here we use an atomistic strategy to elucidate the spectral properties of Mch. purpuratum LH2 and understand the differences with the most commonly studied LH2 from Rbl. acidophilus. Our strategy exploits a combination of molecular dynamics simulations, multiscale polarizable quantum mechanics/molecular mechanics calculations, and lineshape simulations. Our calculations reveal that the spectral properties of LH2 complexes are tuned by site energies and exciton couplings, which in turn depend on the structural fluctuations of the bacteriochlorophylls. Our strategy proves effective in reproducing the absorption and CD spectra of the two LH2 complexes, and in uncovering the origin of their differences. This work proves that it is possible to obtain insight into the spectral tuning strategies of purple bacteria by quantitatively simulating the spectral properties of their antenna complexes.

A bacterium from a mountain lake harvests light using both proton-pumping xanthorhodopsins and bacteriochlorophyll-based photosystems.

Photoheterotrophic bacteria harvest light energy using either proton-pumping rhodopsins or bacteriochlorophyll (BChl)-based photosystems. The bacterium Sphingomonas glacialis AAP5 isolated from the alpine lake Gossenköllesee contains genes for both systems. Here, we show that BChl is expressed between 4°C and 22°C in the dark, whereas xanthorhodopsin is expressed only at temperatures below 16°C and in the presence of light. Thus, cells grown at low temperatures under a natural light-dark cycle contain both BChl-based photosystems and xanthorhodopsins with a nostoxanthin antenna. Flash photolysis measurements proved that both systems are photochemically active. The captured light energy is used for ATP synthesis and stimulates growth. Thus, S. glacialis AAP5 represents a chlorophototrophic and a retinalophototrophic organism. Our analyses suggest that simple xanthorhodopsin may be preferred by the cells under higher light and low temperatures, whereas larger BChl-based photosystems may perform better at lower light intensities. This indicates that the use of two systems for light harvesting may represent an evolutionary adaptation to the specific environmental conditions found in alpine lakes and other analogous ecosystems, allowing bacteria to alternate their light-harvesting machinery in response to large seasonal changes of irradiance and temperature.

Phasor Analysis Reveals Multicomponent Fluorescence Kinetics in the LH2 Complex from Marichromatium purpuratum.

We investigated the fluorescence kinetics of LH2 complexes from Marichromatium purpuratum, the cryo-EM structure of which has been recently elucidated with 2.4 Å resolution. The experiments have been carried out as a function of the excitation density by varying both the excitation fluence and the repetition rate of the laser excitation. Instead of the usual multiexponential fitting procedure, we applied the less common phasor formalism for evaluating the transients because this allows for a model-free analysis of the data without a priori knowledge about the number of processes that contribute to a particular decay. For the various excitation conditions, this analysis reproduces consistently three lifetime components with decay times below 100 ps, 500 ps, and 730 ps, which were associated with the quenched state, singlet-triplet annihilation, and fluorescence decay, respectively. Moreover, it reveals that the number of decay components that contribute to the transients depends on whether the excitation wavelength is in resonance with the B800 BChl a molecules or with the carotenoids. Based on the mutual arrangement of the chromophores in their binding pockets, this leads us to conclude that the energy transfer pathways within the LH2 complex of this species differ significantly from each other for exciting either the B800 BChl molecules or the carotenoids. Finally, we speculate whether the illumination with strong laser light converts the LH2 complexes studied here into a quenched conformation that might be related to the development of the non-photochemical quenching mechanism that occurs in higher plants.

Cryo-EM structures of light-harvesting 2 complexes from Rhodopseudomonas palustris reveal the molecular origin of absorption tuning.

The genomes of some purple photosynthetic bacteria contain a multigene puc family encoding a series of α- and ß-polypeptides that together form a heterogeneous antenna of light-harvesting 2 (LH2) complexes. To unravel this complexity, we generated four sets of puc deletion mutants in Rhodopseudomonas palustris, each encoding a single type of pucBA gene pair and enabling the purification of complexes designated as PucA-LH2, PucB-LH2, PucD-LH2, and PucE-LH2. The structures of all four purified LH2 complexes were determined by cryogenic electron microscopy (cryo-EM) at resolutions ranging from 2.7 to 3.6 Å. Uniquely, each of these complexes contains a hitherto unknown polypeptide, γ, that forms an extended undulating ribbon that lies in the plane of the membrane and that encloses six of the nine LH2 αß-subunits. The γ-subunit, which is located near to the cytoplasmic side of the complex, breaks the C9 symmetry of the LH2 complex and binds six extra bacteriochlorophylls (BChls) that enhance the 800-nm absorption of each complex. The structures show that all four complexes have two complete rings of BChls, conferring absorption bands centered at 800 and 850 nm on the PucA-LH2, PucB-LH2, and PucE-LH2 complexes, but, unusually, the PucD-LH2 antenna has only a single strong near-infared (NIR) absorption peak at 803 nm. Comparison of the cryo-EM structures of these LH2 complexes reveals altered patterns of hydrogen bonds between LH2 αß-side chains and the bacteriochlorin rings, further emphasizing the major role that H bonds play in spectral tuning of bacterial antenna complexes.

Carotenoid responds to excess energy dissipation in the LH2 complex from Rhodoblastus acidophilus.

The functions of both (bacterio) chlorophylls and carotenoids in light-harvesting complexes have been extensively studied during the past decade, yet, the involvement of BChl a high-energy Soret band in the cascade of light-harvesting processes still remains a relatively unexplored topic. Here, we present transient absorption data recorded after excitation of the Soret band in the LH2 complex from Rhodoblastus acidophilus. Comparison of obtained data to those recorded after excitation of rhodopin glucoside and B800 BChl a suggests that no Soret-to-Car energy transfer pathway is active in LH2 complex. Furthermore, a spectrally rich pattern observed in the spectral region of rhodopin glucoside ground state bleaching (420-550 nm) has been assigned to an electrochromic shift. The results of global fitting analysis demonstrate two more features. A 6 ps component obtained exclusively after excitation of the Soret band has been assigned to the response of rhodopin glucoside to excess energy dissipation in LH2. Another time component, ~ 450 ps, appearing independently of the excitation wavelength was assigned to BChl a-to-Car triplet-triplet transfer. Presented data demonstrate several new features of LH2 complex and its behavior following the excitation of the Soret band.

2.4-Å structure of the double-ring Gemmatimonas phototrophica photosystem.

Phototrophic Gemmatimonadetes evolved the ability to use solar energy following horizontal transfer of photosynthesis-related genes from an ancient phototrophic proteobacterium. The electron cryo-microscopy structure of the Gemmatimonas phototrophica photosystem at 2.4 Å reveals a unique, double-ring complex. Two unique membrane-extrinsic polypeptides, RC-S and RC-U, hold the central type 2 reaction center (RC) within an inner 16-subunit light-harvesting 1 (LH1) ring, which is encircled by an outer 24-subunit antenna ring (LHh) that adds light-gathering capacity. Femtosecond kinetics reveal the flow of energy within the RC-dLH complex, from the outer LHh ring to LH1 and then to the RC. This structural and functional study shows that G. phototrophica has independently evolved its own compact, robust, and highly effective architecture for harvesting and trapping solar energy.

Vibrational Modes Promoting Exciton Relaxation in the B850 Band of LH2.

Exciton relaxation dynamics in multichromophore systems are often modeled using Redfield theory, where bath fluctuations mediate the relaxation among the exciton eigenstates. Identifying the vibrational or phonon modes that are implicated in exciton relaxation allows more detailed understanding of exciton dynamics. Here we focus on a well-studied light-harvesting II complex (LH2) isolated from the photosynthetic purple bacterium Rhodoblastus acidophilus strain 10050. Using two synchronized mode-locked lasers, we carried out a polarization-dependent two-dimensional electronic spectroscopy (2DES) study of an ultrafast exciton relaxation in the B850 band of LH2. 2DES data with different polarization configurations enable us to investigate the exciton relaxation between the k = ±1 exciton states. Then, we identify vibrational modes coupled to the exciton relaxation by analyzing the coherent wavepackets in the 2DES signals. Focusing on the coherent vibrational wavepackets, the data suggest that certain symmetry-breaking modes of monomeric units play a key role in exciton relaxation.

Intramolecular charge-transfer enhances energy transfer efficiency in carotenoid-reconstituted light-harvesting 1 complex of purple photosynthetic bacteria.

In bacterial photosynthesis, the excitation energy transfer (EET) from carotenoids to bacteriochlorophyll a has a significant impact on the overall efficiency of the primary photosynthetic process. This efficiency can be enhanced when the involved carotenoid has intramolecular charge-transfer (ICT) character, as found in light-harvesting systems of marine alga and diatoms. Here, we provide insights into the significance of ICT excited states following the incorporation of a higher plant carotenoid, ß-apo-8'-carotenal, into the carotenoidless light-harvesting 1 (LH1) complex of the purple photosynthetic bacterium Rhodospirillum rubrum strain G9+. ß-apo-8'-carotenal generates the ICT excited state in the reconstituted LH1 complex, achieving an efficiency of EET of up to 79%, which exceeds that found in the wild-type LH1 complex.

Structure elucidation of the novel carotenoid gemmatoxanthin from the photosynthetic complex of Gemmatimonas phototrophica AP64.

Gemmatimonas phototrophica AP64 is the first phototrophic representative of the bacterial phylum Gemmatimonadetes. The cells contain photosynthetic complexes with bacteriochlorophyll a as the main light-harvesting pigment and an unknown carotenoid with a single broad absorption band at 490 nm in methanol. The carotenoid was extracted from isolated photosynthetic complexes, and purified by liquid chromatography. A combination of nuclear magnetic resonance (1H NMR, COSY, 1H-13C HSQC, 1H-13C HMBC, J-resolved, and ROESY), high-resolution mass spectroscopy, Fourier-transformed infra-red, and Raman spectroscopy was used to determine its chemical structure. The novel linear carotenoid, that we have named gemmatoxanthin, contains 11 conjugated double bonds and is further substituted by methoxy, carboxyl and aldehyde groups. Its IUPAC-IUBMB semi-systematic name is 1'-Methoxy-19'-oxo-3',4'-didehydro-7,8,1',2'-tetrahydro- Ψ, Ψ carotene-16-oic acid. To our best knowledge, the presence of the carboxyl, methoxy and aldehyde groups on a linear C40 carotenoid backbone is reported here for the first time.

Low-Frequency Vibronic Mixing Modulates the Excitation Energy Flow in Bacterial Light-Harvesting Complex II.

Oscillatory features observed in two-dimensional electronic spectroscopy (2DES) manifest coherent vibrational and electronic dynamics and even the interplay of them. Recently, we developed a 2DES technique utilizing a pair of synchronized repetition-frequency-stabilized lasers, which enables the wide dynamic range measurements of 2DES signals rapidly. Here, we apply this dual-laser 2DES technique to investigate the electronic energy transfer (EET) process in bacterial light-harvesting complex II consisting of B800 and B850 circular aggregates at ambient temperature, and the coherent vibrational wavepakcet associated with the EET between the two aggregates are measured. Examining the principal component analysis of the time-resolved 2DES signal, we found that the EET from B800 to low-lying B850 states is modulated by a low-frequency (156 cm-1) vibrational mode of the exciton donor (B800). This observation suggests that the donor transition density is modulated by this vibration, which, in turn, modulates the energy transfer dynamics.

Time-Domain Line-Shape Analysis from 2D Spectroscopy to Precisely Determine Hamiltonian Parameters for a Photosynthetic Complex.

Optical signals come from coherences between quantum states, with spectral line widths determined by the coherences' dephasing dynamics. Using a 2D electronic spectrometer, we observe weak coherence- and rephasing-time-domain signals persisting to 1 ps in the Fenna-Matthews-Olson complex at 77 K. These are coherences between the ground and excited states prepared after the complex interacts once or three times with light, rather than zero-quantum coherences that are more frequently investigated following two interactions. Here, we use these small but persistent signal components to isolate spectral contributions with narrowed peaks and reveal the system's eigenenergies.

The 2.4 Å cryo-EM structure of a heptameric light-harvesting 2 complex reveals two carotenoid energy transfer pathways.

We report the 2.4 Ångström resolution structure of the light-harvesting 2 (LH2) complex from Marichromatium (Mch.) purpuratum determined by cryogenic electron microscopy. The structure contains a heptameric ring that is unique among all known LH2 structures, explaining the unusual spectroscopic properties of this bacterial antenna complex. We identify two sets of distinct carotenoids in the structure and describe a network of energy transfer pathways from the carotenoids to bacteriochlorophyll a molecules. The geometry imposed by the heptameric ring controls the resonant coupling of the long-wavelength energy absorption band. Together, these details reveal key aspects of the assembly and oligomeric form of purple bacterial LH2 complexes that were previously inaccessible by any technique.

A comparative look at structural variation among RC-LH1 'Core' complexes present in anoxygenic phototrophic bacteria.

All purple photosynthetic bacteria contain RC-LH1 'Core' complexes. The structure of this complex from Rhodobacter sphaeroides, Rhodopseudomonas palustris and Thermochromatium tepidum has been solved using X-ray crystallography. Recently, the application of single particle cryo-EM has revolutionised structural biology and the structure of the RC-LH1 'Core' complex from Blastochloris viridis has been solved using this technique, as well as the complex from the non-purple Chloroflexi species, Roseiflexus castenholzii. It is apparent that these structures are variations on a theme, although with a greater degree of structural diversity within them than previously thought. Furthermore, it has recently been discovered that the only phototrophic representative from the phylum Gemmatimonadetes, Gemmatimonas phototrophica, also contains a RC-LH1 'Core' complex. At present only a low-resolution EM-projection map exists but this shows that the Gemmatimonas phototrophica complex contains a double LH1 ring. This short review compares these different structures and looks at the functional significance of these variations from two main standpoints: energy transfer and quinone exchange.

Gemmatimonas groenlandica sp. nov. Is an Aerobic Anoxygenic Phototroph in the Phylum Gemmatimonadetes.

The bacterial phylum Gemmatimonadetes contains members capable of performing bacteriochlorophyll-based phototrophy (chlorophototrophy). However, only one strain of chlorophototrophic Gemmatimonadetes bacteria (CGB) has been isolated to date, hampering our further understanding of their photoheterotrophic lifestyle and the evolution of phototrophy in CGB. By combining a culturomics strategy with a rapid screening technique for chlorophototrophs, we report the isolation of a new member of CGB, Gemmatimonas (G.) groenlandica sp. nov., from the surface water of a stream in the Zackenberg Valley in High Arctic Greenland. Distinct from the microaerophilic G. phototrophica strain AP64T, G. groenlandica strain TET16T is a strictly aerobic anoxygenic phototroph, lacking many oxygen-independent enzymes while possessing an expanded arsenal for coping with oxidative stresses. Its pigment composition and infra-red absorption properties are also different from G. phototrophica, indicating that it possesses a different photosystem apparatus. The complete genome sequence of G. groenlandica reveals unique and conserved features in the photosynthesis gene clusters of CGB. We further analyzed metagenome-assembled genomes of CGB obtained from soil and glacier metagenomes from Northeast Greenland, revealing a wide distribution pattern of CGB beyond the stream water investigated.

Before Förster. Initial excitation in photosynthetic light harvesting.

Electronic 2D spectroscopy allows nontrivial quantum effects to be explored in unprecedented detail. Here, we apply recently developed fluorescence detected coherent 2D spectroscopy to study the light harvesting antenna 2 (LH2) of photosynthetic purple bacteria. We report double quantum coherence 2D spectra which show clear cross peaks indicating correlated excitations. Similar results are found for rephasing and nonrephasing signals. Analysis of signal generating quantum pathways leads to the conclusion that, contrary to the currently prevailing physical picture, the two weakly coupled pigment rings of LH2 share the initial electronic excitation leading to quantum mechanical correlation between the two clearly separate absorption bands. These results are general and have consequences for the interpretation of initially created excited states not only in photosynthesis but in all light absorbing systems composed of weakly interacting pigments where the excitation transfer is commonly described by using Förster theory. Being able to spectrally resolve the nonequilibrium dynamics immediately following photoabsorption may provide a glimpse to the systems' transition into the Förster regime.

Assessing density functional theory in real-time and real-space as a tool for studying bacteriochlorophylls and the light-harvesting complex 2.

We use real-time density functional theory on a real-space grid to calculate electronic excitations of bacteriochlorophyll chromophores of the light-harvesting complex 2 (LH2). Comparison with Gaussian basis set calculations allows us to assess the numerical trust range for computing electron dynamics in coupled chromophores with both types of techniques. Tuned range-separated hybrid calculations for one bacteriochlorophyll as well as two coupled ones are used as a reference against which we compare results from the adiabatic time-dependent local density approximation (TDLDA). The tuned range-separated hybrid calculations lead to a qualitatively correct description of the electronic excitations and couplings. They allow us to identify spurious charge-transfer excitations that are obtained with the TDLDA. When we take into account the environment that the LH2 protein complex forms for the bacteriochlorophylls, we find that it substantially shifts the energy of the spurious charge-transfer excitations, restoring a qualitatively correct electronic coupling of the dominant excitations also for TDLDA.