ApcE plays an important role in light-induced excitation energy dissipation in the Synechocystis PCC6803 phycobilisomes.

Phycobilisomes (PBs) play an important role in cyanobacterial photosynthesis. They capture light and transfer excitation energy to the photosynthetic reaction centres. PBs are also central to some photoprotective and photoregulatory mechanisms that help sustain photosynthesis under non-optimal conditions. Amongst the mechanisms involved in excitation energy dissipation that are activated in response to excessive illumination is a recently discovered light-induced mechanism that is intrinsic to PBs and has been the least studied. Here, we used single-molecule spectroscopy and developed robust data analysis methods to explore the role of a terminal emitter subunit, ApcE, in this intrinsic, light-induced mechanism. We isolated the PBs from WT Synechocystis PCC 6803 as well as from the ApcE-C190S mutant of this strain and compared the dynamics of their fluorescence emission. PBs isolated from the mutant (i.e., ApcE-C190S-PBs), despite not binding some of the red-shifted pigments in the complex, showed similar global emission dynamics to WT-PBs. However, a detailed analysis of dynamics in the core revealed that the ApcE-C190S-PBs are less likely than WT-PBs to enter quenched states under illumination but still fully capable of doing so. This result points to an important but not exclusive role of the ApcE pigments in the light-induced intrinsic excitation energy dissipation mechanism in PBs.

Investigation into aggregation types in a benzodithiophene-isoindigo copolymer.

The type of aggregation in conjugated polymers determines their use in electronic devices. H-type aggregates are most suitable for solar cell applications, while J-type aggregates are recommended for light-emitting diodes. In this work, we used three methods to determine the type of aggregates in a benzodithiophene-isoindigo-based (PBDTI-DT) copolymer, namely, Huang-Rhys factor evolution with temperature, Franck-Condon analysis, and relative quantum yield (QY) calculation. All three methods indicate that both aggregation types are present, and the QY calculation clearly indicates that H-aggregates are more dominant. Time-dependent density functional theory was used to identify the two absorption bands of PBDTI-DT as local π - π* and intramolecular charge-transfer transitions.

Theoretical comparison of real-time feedback-driven single-particle tracking techniques.

Real-time feedback-driven single-particle tracking is a technique that uses feedback control to enable single-molecule spectroscopy of freely diffusing particles in native or near-native environments. A number of different real-time feedback-driven single-particle tracking (RT-FD-SPT) approaches exist, and comparisons between methods based on experimental results are of limited use due to differences in samples and setups. In this study, we used statistical calculations and dynamical simulations to directly compare the performance of different methods. The methods considered were the orbital method, the knight's tour (grid scan) method, and MINFLUX, and we considered both fluorescence-based and interferometric scattering (iSCAT) approaches. There is a fundamental trade-off between precision and speed, with the knight's tour method being able to track the fastest diffusion but with low precision, and MINFLUX being the most precise but only tracking slow diffusion. To compare iSCAT and fluorescence, different biological samples were considered, including labeled and intrinsically fluorescent samples. The success of iSCAT as compared to fluorescence is strongly dependent on the particle size and the density and photophysical properties of the fluorescent particles. Using a wavelength for iSCAT that is negligibly absorbed by the tracked particle allows for an increased illumination intensity, which results in iSCAT providing better tracking for most samples. This work highlights the fundamental aspects of performance in RT-FD-SPT and should assist with the selection of an appropriate method for a particular application. The approach used can easily be extended to other RT-FD-SPT methods.

Real-Time Feedback-Driven Single-Particle Tracking: A Survey and Perspective.

Real-time feedback-driven single-particle tracking (RT-FD-SPT) is a class of techniques in the field of single-particle tracking that uses feedback control to keep a particle of interest in a detection volume. These methods provide high spatiotemporal resolution on particle dynamics and allow for concurrent spectroscopic measurements. This review article begins with a survey of existing techniques and of applications where RT-FD-SPT has played an important role. Each of the core components of RT-FD-SPT are systematically discussed in order to develop an understanding of the trade-offs that must be made in algorithm design and to create a clear picture of the important differences, advantages, and drawbacks of existing approaches. These components are feedback tracking and control, ranging from simple proportional-integral-derivative control to advanced nonlinear techniques, estimation to determine particle location from the measured data, including both online and offline algorithms, and techniques for calibrating and characterizing different RT-FD-SPT methods. Then a collection of metrics for RT-FD-SPT is introduced to help guide experimentalists in selecting a method for their particular application and to help reveal where there are gaps in the techniques that represent opportunities for further development. Finally, this review is concluded with a discussion on future perspectives in the field.

Mono-Doped and Co-Doped Nanostructured Hematite for Improved Photoelectrochemical Water Splitting.

In this study, zinc-doped (α-Fe2O3:Zn), silver-doped (α-Fe2O3:Ag) and zinc/silver co-doped hematite (α-Fe2O3:Zn/Ag) nanostructures were synthesized by spray pyrolysis. The synthesized nanostructures were used as photoanodes in the photoelectrochemical (PEC) cell for water-splitting. A significant improvement in photocurrent density of 0.470 mAcm-2 at 1.23 V vs. reversible hydrogen electrode (RHE) was recorded for α-Fe2O3:Zn/Ag. The α-Fe2O3:Ag, α-Fe2O3:Zn and pristine hematite samples produced photocurrent densities of 0.270, 0.160, and 0.033 mAcm-2, respectively. Mott-Schottky analysis showed that α-Fe2O3:Zn/Ag had the highest free carrier density of 8.75 × 1020 cm-3, while pristine α-Fe2O3, α-Fe2O3:Zn, α-Fe2O3:Ag had carrier densities of 1.57 × 1019, 5.63 × 1020, and 6.91 × 1020 cm-3, respectively. Electrochemical impedance spectra revealed a low impedance for α-Fe2O3:Zn/Ag. X-ray diffraction confirmed the rhombohedral corundum structure of hematite. Scanning electron microscopy micrographs, on the other hand, showed uniformly distributed grains with an average size of <30 nm. The films were absorbing in the visible region with an absorption onset ranging from 652 to 590 nm, corresponding to a bandgap range of 1.9 to 2.1 eV. Global analysis of ultrafast transient absorption spectroscopy data revealed four decay lifetimes, with a reduction in the electron-hole recombination rate of the doped samples on a timescale of tens of picoseconds.

What to feed or what not to feed-that is still the question.

INTRODUCTION: This review addresses metabolic diversities after grain feeding of cattle using artificial total mixed ration (TMR), in place of pasture-based feeding. OBJECTIVES: To determine how grain feeding impairs the deuterium-depleting functions of the anaplerotic mitochondrial matrix during milk and meat production. METHODS: Based on published data we herein evaluate how grain-fed animals essentially follow a branched-chain amino acid and odd-chain fatty acid-based reductive carboxylation-dependent feedstock, which is also one of the mitochondrial deuterium-accumulating dysfunctions in human cancer. RESULTS: It is now evident that food-based intracellular deuterium exchange reactions, especially that of glycogenic substrate oxidation, are significant sources of deuterium-enriched (2H; D) metabolic water with a significant impact on animal and human health. The burning of high deuterium nutritional dairy products into metabolic water upon oxidation in the human body may contribute to similar metabolic conditions and diseases as described in state-of-the-art articles for cows. Grain feeding also limits oxygen delivery to mitochondria for efficient deuterium-depleted metabolic water production by glyphosate herbicide exposure used in genetically modified crops of TMR constituents. CONCLUSION: Developments in medical metabolomics, biochemistry and deutenomics, which is the science of biological deuterium fractionation and discrimination warrant urgent critical reviews in order to control the epidemiological scale of population diseases such as diabetes, obesity and cancer by a thorough understanding of how the compromised metabolic health of grain-fed dairy cows impacts human consumers.

Sub-lithic photosynthesis in hot desert habitats.

In hyper-arid soil environments, photosynthetic microorganisms are largely restricted to hypolithic (sub-lithic) habitats: i.e., on the ventral surfaces of translucent pebbles in desert pavements. Here, we combined fluorometric, spectroscopic, biochemical and metagenomic approaches to investigate in situ the light transmission properties of quartz stones in the Namib Desert, and assess the photosynthetic activity of the underlying hypolithic cyanobacterial biofilms. Quartz pebbles greatly reduced the total photon flux to the ventral surface biofilms and filtered out primarily the short wavelength portion of the solar spectrum. Chlorophylls d and f were not detected in biofilm pigment extracts; however, hypolithic cyanobacterial communities showed some evidence of adaptation to sub-lithic conditions, including the prevalence of genes encoding Helical Carotenoid Proteins, which are associated with desiccation stress. Under water-saturated conditions, hypolithic communities showed no evidence of light stress, even when the quartz stones were exposed to full midday sunlight. This initial study creates a foundation for future in-situ and laboratory exploration of various adaptation mechanisms employed by photosynthetic organisms forming hypolithic microbial communities.

Photoelectrochemical performance and ultrafast dynamics of photogenerated electrons and holes in highly titanium-doped hematite.

Elemental doping of hematite has been widely performed to improve its mobility, electrical conductivity as well as to suppress electron-hole recombination in photoelectrochemical applications. When hematite is doped with high titanium concentrations, above 5%, pseudobrookite layers may be formed as overlayers leading to improved photocurrent while further doping beyond 15% could lead to the formation of a titania overlayer which has an effect of suppressing photocurrent. In this study, we observed that doping hematite with titanium improves photocurrent, reaching a maximum of 1.83 mA cm-2 at a titanium concentration of 15%, the highest achieved photocurrent with spin coating method. Further titanium incorporation to 20% resulted in a decrease of the photocurrent. XRD measurements shows that a Fe2TiO5 layer formed at 15% Ti concentration which resulted in the observed increase in photocurrent while a reduction in photocurrent at 20% Ti concentration could have resulted from the formation of a TiO2 layer. Analysis of the transient absorption spectroscopy data was achieved using a four-component sequential analysis scheme in the Glotaran software. We observed major doping concentration dependent lifetimes in the τ3 and τ4 values where the 15% doped samples had the slowest recombination rates. We also observed a blueshift in the spectra with increasing doping concentration, suggesting the occurance of the Burstein-Moss effect. This work shows that doping hematite with titanium leads to structural changes of the photoanodes at Ti concentrations of over 10%, in addition to the well documented conductivity enhancement.

Charge transfer states in phycobilisomes.

Phycobilisomes (PBs) absorb light and supply downstream photosynthetic processes with excitation energy in many cyanobacteria and algae. In response to a sudden increase in light intensity, excess excitation energy is photoprotectively dissipated in PBs by means of the orange carotenoid protein (OCP)-related mechanism or via a light-activated intrinsic decay channel. Recently, we have identified that both mechanisms are associated with far-red emission states. Here, we investigate the far-red states involved with the light-induced intrinsic mechanism by exploring the energy landscape and electro-optical properties of the pigments in PBs. While Stark spectroscopy showed that the far-red states in PBs exhibit a strong charge-transfer (CT) character at cryogenic temperatures, single molecule spectroscopy revealed that CT states should also be present at room temperature. Owing to the strong environmental sensitivity of CT states, the knowledge gained from this study may contribute to the design of a new generation of fluorescence markers.

Strong plasmonic fluorescence enhancement of individual plant light-harvesting complexes.

Plasmonic coupling of metallic nanoparticles and adjacent pigments can dramatically increase the brightness of the pigments due to the enhanced local electric field. Here, we demonstrate that the fluorescence brightness of a single plant light-harvesting complex (LHCII) can be significantly enhanced when coupled to a gold nanorod (AuNR). The AuNRs utilized in this study were prepared via chemical reactions, and the hybrid system was constructed using a simple and economical spin-assisted layer-by-layer technique. Enhancement of fluorescence brightness of up to 240-fold was observed, accompanied by a 109-fold decrease in the average (amplitude-weighted) fluorescence lifetime from approximately 3.5 ns down to 32 ps, corresponding to an excitation enhancement of 63-fold and emission enhancement of up to 3.8-fold. This large enhancement is due to the strong spectral overlap of the longitudinal localized surface plasmon resonance of the utilized AuNRs and the absorption or emission bands of LHCII. This study provides an inexpensive strategy to explore the fluorescence dynamics of weakly emitting photosynthetic light-harvesting complexes at the single molecule level.

The role of far-red spectral states in the energy regulation of phycobilisomes.

The main light-harvesting pigment-protein complex of cyanobacteria and certain algae is the phycobilisome, which harvests sunlight and regulates the flow of absorbed energy to provide the photochemical reaction centres with a constant energy throughput. At least two light-driven mechanisms of excited energy quenching in phycobilisomes have been identified: the dominant mechanism in many strains of cyanobacteria depends on the orange carotenoid protein (OCP), while the second mechanism is intrinsically available to a phycobilisome and is possibly activated faster than the former. Recent single molecule spectroscopy studies have shown that far-red (FR) emission states are related to the OCP-dependent mechanism and it was proposed that the second mechanism may involve similar states. In this study, we examined the dynamics of simultaneously measured emission spectra and intensities from a large set of individual phycobilisome complexes from Synechocystis PCC 6803. Our results suggest a direct relationship between FR spectral states and thermal energy dissipating states and can be explained by a single phycobilin pigment in the phycobilisome core acting as the site of both quenching and FR emission likely due to the presence of a charge-transfer state. Our experimental method provides a means to accurately resolve the fluorescence lifetimes and spectra of the FR states, which enabled us to quantify a kinetic model that reproduces most of the experimentally determined properties of the FR states.

The future of quantum biology.

Biological systems are dynamical, constantly exchanging energy and matter with the environment in order to maintain the non-equilibrium state synonymous with living. Developments in observational techniques have allowed us to study biological dynamics on increasingly small scales. Such studies have revealed evidence of quantum mechanical effects, which cannot be accounted for by classical physics, in a range of biological processes. Quantum biology is the study of such processes, and here we provide an outline of the current state of the field, as well as insights into future directions.

Energy dissipation mechanisms in the FCPb light-harvesting complex of the diatom Cyclotella meneghiniana.

Transient absorption spectroscopy has been applied to investigate the energy dissipation mechanisms in the nonameric fucoxanthin-chlorophyll-a,c-binding protein FCPb of the centric diatom Cyclotella meneghiniana. FCPb complexes in their unquenched state were compared with those in two types of quenching environments, namely aggregation-induced quenching by detergent removal, and clustering via incorporation into liposomes. Applying global and target analysis, in combination with a fluorescence lifetime study and annihilation calculations, we were able to resolve two quenching channels in FCPb that involve chlorophyll-a pigments for FCPb exposed to both quenching environments. The fast quenching channel operates on a timescale of tens of picoseconds and exhibits similar spectral signatures as the unquenched state. The slower quenching channel operates on a timescale of tens to hundreds of picoseconds, depending on the degree of quenching, and is characterized by enhanced population of low-energy states between 680 and 710 nm. The results indicate that FCPb is, in principle, able to function as a dissipater of excess energy and can do this in vitro even more efficiently than the homologous FCPa complex, the sole complex involved in fast photoprotection in these organisms. This indicates that when a complex displays photoprotection-related spectral signatures in vitro it does not imply that the complex participates in photoprotection in vivo. We suggest that FCPa is favored over FCPb as the sole energy-regulating complex in diatoms because its composition can more easily establish the balance between light-harvesting and quenching required for efficient photoprotection.

Orthonormal vector general polynomials derived from the Cartesian gradient of the orthonormal Zernike-based polynomials.

The concept of orthonormal vector circle polynomials is revisited by deriving a set from the Cartesian gradient of Zernike polynomials in a unit circle using a matrix-based approach. The heart of this model is a closed-form matrix equation of the gradient of Zernike circle polynomials expressed as a linear combination of lower-order Zernike circle polynomials related through a gradient matrix. This is a sparse matrix whose elements are two-dimensional standard basis transverse Euclidean vectors. Using the outer product form of the Cholesky decomposition, the gradient matrix is used to calculate a new matrix, which we used to express the Cartesian gradient of the Zernike circle polynomials as a linear combination of orthonormal vector circle polynomials. Since this new matrix is singular, the orthonormal vector polynomials are recovered by reducing the matrix to its row echelon form using the Gauss-Jordan elimination method. We extend the model to derive orthonormal vector general polynomials, which are orthonormal in a general pupil by performing a similarity transformation on the gradient matrix to give its equivalent in the general pupil. The outer form of the Gram-Schmidt procedure and the Gauss-Jordan elimination method are then applied to the general pupil to generate the orthonormal vector general polynomials from the gradient of the orthonormal Zernike-based polynomials. The performance of the model is demonstrated with a simulated wavefront in a square pupil inscribed in a unit circle.

Switching an Individual Phycobilisome Off and On.

Photosynthetic organisms have found various smart ways to cope with unexpected changes in light conditions. In many cyanobacteria, the lethal effects of a sudden increase in light intensity are mitigated mainly by the interaction between phycobilisomes (PBs) and the orange carotenoid protein (OCP). The latter senses high light intensities by means of photoactivation and triggers thermal energy dissipation from the PBs. Due to the brightness of their emission, PBs can be characterized at the level of individual complexes. Here, energy dissipation from individual PBs was reversibly switched on and off using only light and OCP. We reveal the presence of quasistable intermediate states during the binding and unbinding of OCP to PB, with a spectroscopic signature indicative of transient decoupling of some of the PB rods during docking of OCP. Real-time control of emission from individual PBs has the potential to contribute to the development of new super-resolution imaging techniques.

Phycocyanin: One Complex, Two States, Two Functions.

Solar energy captured by pigments embedded in light-harvesting complexes can be transferred to neighboring pigments, dissipated, or emitted as fluorescence. Only when it reaches a reaction center is the excitation energy stabilized in the form of a charge separation and converted into chemical energy. Well-directed and regulated energy transfer within the network of pigments is therefore of crucial importance for the success of the photosynthetic processes. Using single-molecule spectroscopy, we show that phycocyanin can dynamically switch between two spectrally distinct states originating from two different conformations. Unexpectedly, one of the two states has a red-shifted emission spectrum. This state is not involved in energy dissipation; instead, we propose that it is involved in direct energy transfer to photosystem I. Finally, our findings suggest that the function of linker proteins in phycobilisomes is to stabilize one state or the other, thus controlling the light-harvesting functions of phycocyanin.

Apoprotein heterogeneity increases spectral disorder and a step-wise modification of the B850 fluorescence peak position.

It has already been established that the quaternary structure of the main light-harvesting complex (LH2) from the photosynthetic bacterium Rhodopseudomonas palustris is a nonameric 'ring' of PucAB heterodimers and under low-light culturing conditions an increased diversity of PucB synthesis occurs. In this work, single molecule fluorescence emission studies show that different classes of LH2 'rings' are present in "low-light" adapted cells and that an unknown chaperon process creates multiple sub-types of 'rings' with more conformational sub-states and configurations. This increase in spectral disorder significantly augments the cross-section for photon absorption and subsequent energy flow to the reaction centre trap when photon availability is a limiting factor. This work highlights yet another variant used by phototrophs to gather energy for cellular development.

How reduced excitonic coupling enhances light harvesting in the main photosynthetic antennae of diatoms.

Strong excitonic interactions are a key design strategy in photosynthetic light harvesting, expanding the spectral cross-section for light absorption and creating considerably faster and more robust excitation energy transfer. These molecular excitons are a direct result of exceptionally densely packed pigments in photosynthetic proteins. The main light-harvesting complexes of diatoms, known as fucoxanthin-chlorophyll proteins (FCPs), are an exception, displaying surprisingly weak excitonic coupling between their chlorophyll (Chl) a's, despite a high pigment density. Here, we show, using single-molecule spectroscopy, that the FCP complexes of Cyclotella meneghiniana switch frequently into stable, strongly emissive states shifted 4-10 nm toward the red. A few percent of isolated FCPa complexes and ∼20% of isolated FCPb complexes, on average, were observed to populate these previously unobserved states, percentages that agree with the steady-state fluorescence spectra of FCP ensembles. Thus, the complexes use their enhanced sensitivity to static disorder to increase their light-harvesting capability in a number of ways. A disordered exciton model based on the structure of the main plant light-harvesting complex explains the red-shifted emission by strong localization of the excitation energy on a single Chl a pigment in the terminal emitter domain due to very specific pigment orientations. We suggest that the specific construction of FCP gives the complex a unique strategy to ensure that its light-harvesting function remains robust in the fluctuating protein environment despite limited excitonic interactions.

Strehl ratio and amplitude-weighted generalized orthonormal Zernike-based polynomials.

The concept of orthonormal polynomials is revisited by developing a Zernike-based orthonormal set for a non-circular pupil that is transmitting an aberrated, non-uniform field. We refer to this pupil as a general pupil. The process is achieved by using the matrix form of the Gram-Schmidt procedure on Zernike circle polynomials and is interpreted as a process of balancing each Zernike circle polynomial by adding those of lower order in the general pupil, a procedure which was previously performed using classical aberrations. We numerically demonstrate this concept by comparing the representation of phase in a square-Gaussian pupil using the Zernike-Gauss square and Zernike circle polynomials. As expected, using the Strehl ratio, we show that only specific lower-order aberrations can be used to balance specific aberrations, for example, tilt cannot be used to balance spherical aberration. In the process, we present a possible definition of the Maréchal criterion for the analysis of the tolerance of systems with apodized pupils.