Jianzhong Wen

Long-lived quantum coherence in photosynthetic complexes at physiological temperature

Photosynthetic antenna complexes capture and concentrate solar radiation by transferring the excitation to the reaction center that stores energy from the photon in chemical bonds. This process occurs with near-perfect quantum efficiency. Recent experiments at cryogenic temperatures have revealed that coherent energy transfer—a wave-like transfer mechanism—occurs in many photosynthetic pigment-protein complexes. Using the Fenna–Matthews–Olson antenna complex (FMO) as a model system, theoretical studies incorporating both incoherent and coherent transfer as well as thermal dephasing predict that environmentally assisted quantum transfer efficiency peaks near physiological temperature; these studies also show that this mechanism simultaneously improves the robustness of the energy transfer process. This theory requires long-lived quantum coherence at room temperature, which never has been observed in FMO. Here we present evidence that quantum coherence survives in FMO at physiological temperature for at least 300 fs, long enough to impact biological energy transport. These data prove that the wave-like energy transfer process discovered at 77 K is directly relevant to biological function. Microscopically, we attribute this long coherence lifetime to correlated motions within the protein matrix encapsulating the chromophores, and we find that the degree of protection afforded by the protein appears constant between 77 K and 277 K. The protein shapes the energy landscape and mediates an efficient energy transfer despite thermal fluctuations.

Membrane orientation of the FMO antenna protein from Chlorobaculum tepidum as determined by mass spectrometry-based footprinting

The high excitation energy-transfer efficiency demanded in photosynthetic organisms relies on the optimal pigment-protein binding orientation in the individual protein complexes and also on the overall architecture of the photosystem. In green sulfur bacteria, the membrane-attached Fenna-Matthews-Olson (FMO) antenna protein functions as a “wire” to connect the large peripheral chlorosome antenna complex with the reaction center (RC), which is embedded in the cytoplasmic membrane (CM). Energy collected by the chlorosome is funneled through the FMO to the RC. Although there has been considerable effort to understand the relationships between structure and function of the individual isolated complexes, the specific architecture for in vivo interactions of the FMO protein, the CM, and the chlorosome, ensuring highly efficient energy transfer, is still not established experimentally. Here, we describe a mass spectrometry-based method that probes solvent-exposed surfaces of the FMO by labeling solvent-exposed aspartic and glutamic acid residues. The locations and extents of labeling of FMO on the native membrane in comparison with it alone and on a chlorosome-depleted membrane reveal the orientation. The large differences in the modification of certain peptides show that the Bchl a #3 side of the FMO trimer interacts with the CM, which is consistent with recent theoretical predictions. Moreover, the results also provide direct experimental evidence to confirm the overall architecture of the photosystem from Chlorobaculum tepidum (C. tepidum) and give information on the packing of the FMO protein in its native environment.

Native Electrospray and Electron-Capture Dissociation in FTICR Mass Spectrometry Provide Top-Down Sequencing of a Protein Component in an Intact Protein Assembly

The intact yeast alcohol dehydrogenase (ADH) tetramer of 147 kDa was introduced into a FTICR mass spectrometer by native electrospray. Electron capture dissociation of the entire 23+ to 27+ charge state distribution produced the expected charge-reduced ions and, more unexpectedly, 39 c-type peptide fragments that identified N-terminus acetylation and the first 55 amino acids. The results are in accord with the crystal structure of yeast ADH, which shows that the C-terminus is buried at the assembly interface, whereas the N-terminus is exposed, allowing ECD to occur. This remarkable observation shows promise that a top-down approach for intact protein assemblies will be effective for characterizing their components, inferring their interfaces, and obtaining both proteomics and structural biology information in one experiment.

Visualization of Excitonic Structure in the Fenna-Matthews-Olson Photosynthetic Complex by Polarization-Dependent Two-Dimensional Electronic Spectroscopy

Photosynthetic light-harvesting proceeds by the collection and highly efficient transfer of energy through a network of pigment-protein complexes. Interchromophore electronic couplings and interactions between pigments and the surrounding protein determine energy levels of excitonic states, and dictate the mechanism of energy flow. The excitonic structure (orientation of excitonic transition dipoles) of pigment-protein complexes is generally deduced indirectly from x-ray crystallography, in combination with predictions of transition energies and couplings in the chromophore site basis. We demonstrate that coarse-grained, excitonic, structural information in the form of projection angles between transition dipole moments can be obtained from the polarization-dependent, two-dimensional electronic spectroscopy of an isotropic sample, particularly when the nonrephasing or free polarization decay signal, rather than the photon echo signal, is considered. This method provides an experimental link between atomic and electronic structure, and accesses dynamical information with femtosecond time resolution. In an investigation of the Fenna-Matthews-Olson complex from green sulfur bacteria, the energy transfer connecting two particular exciton states in the protein was isolated as the primary contributor to a crosspeak in the nonrephasing two-dimensional spectrum at 400 femtoseconds under a specific sequence of polarized excitation pulses. The results suggest the possibility of designing experiments using combinations of tailored polarization sequences to separate and monitor individual relaxation pathways.

Native Electrospray Mass Spectrometry Reveals the Nature and Stoichiometry of Pigments in the FMO Photosynthetic Antenna Protein

The nature and stoichiometry of pigments in the Fenna-Matthews-Olson (FMO) photosynthetic antenna protein complex were determined by native electrospray mass spectrometry. The FMO antenna complex was the first chlorophyll-containing protein that was crystallized. Previous results indicate that the FMO protein forms a trimer with seven bacteriochlorophyll a in each monomer. This model has long been a working basis to understand the molecular mechanism of energy transfer through pigment/pigment and pigment/protein coupling. Recent results have suggested, however, that an eighth bacteriochlorophyll is present in some subunits. In this report, a direct mass spectrometry measurement of the molecular weight of the intact FMO protein complex clearly indicates the existence of an eighth pigment, which is assigned as a bacteriochlorophyll a by mass analysis of the complex and HPLC analysis of the pigment. The eighth pigment is found to be easily lost during purification, which results in its partial occupancy in the mass spectra of the intact complex prepared by different procedures. The results are consistent with the recent X-ray structural models. The existence of the eighth bacteriochlorophyll a in this model antenna protein gives new insights into the functional role of the FMO protein and motivates the need for new theoretical and spectroscopic assignments of spectral features of the FMO protein.

Dynamics of electronic dephasing in the Fenna–Matthews–Olson complex

Electronic coherence has been shown to persist in the Fenna?Matthews?Olson (FMO) antenna complex from green sulfur bacteria at 77?K for at least 660?fs, several times longer than the typical lifetime of a coherence in a dynamic environment at this temperature. Such long-lived coherence was proposed to improve energy transfer efficiency in photosynthetic systems by allowing an excitation to follow a quantum random walk as it approaches the reaction centre. Here we present a model for bath-induced electronic transitions, demonstrating that the protein matrix protects coherences by globally correlating fluctuations in transition energies. We also quantify the dephasing rates for two particular electronic coherences in the FMO complex at 77?K using two-dimensional Fourier transform electronic spectroscopy and find that the lifetimes of individual coherences are distinct. Within the framework of noise-assisted transport, this result suggests that the FMO complex has been locally tuned by natural selection to optimize transfer efficiency by exploiting quantum coherence.

Hydrogen-deuterium exchange mass spectrometry reveals the interaction of Fenna-Matthews-Olson protein and chlorosome CsmA protein.

In green-sulfur bacterial photosynthesis, excitation energy absorbed by a peripheral antenna structure known as the chlorosome is sequentially transferred through a baseplate protein to the Fenna-Matthews-Olson (FMO) antenna protein and into the reaction center, which is embedded in the cytoplasmic membrane. The molecular details of the optimized photosystem architecture required for efficient energy transfer are only partially understood. We address here the question of how the baseplate interacts with the FMO protein by applying hydrogen/deuterium exchange coupled with enzymatic digestion and mass spectrometry analysis to reveal the binding interface of the FMO antenna protein and the CsmA baseplate protein. Several regions on the FMO protein, represented by peptides consisting of 123-129, 140-149, 150-162, 191-208, and 224-232, show significant decreases of deuterium uptake after CsmA binding. The results indicate that the CsmA protein interacts with the Bchl a #1 side of the FMO protein. A global picture including peptide-level details for the architecture of the photosystem from green-sulfur bacteria can now be drawn.

Role of the AcsF Protein in Chloroflexus aurantiacus

The green phototrophic bacteria contain a unique complement of chlorophyll pigments, which self-assemble efficiently into antenna structures known as chlorosomes with little involvement of protein. The few proteins found in chlorosomes have previously been thought to have a primarily structural function. The biosynthetic pathway of the chlorosome pigments, bacteriochlorophylls c, d, and e, is not well understood. In this report, we used spectroscopic, proteomic, and gene expression approaches to investigate the chlorosome proteins of the green filamentous anoxygenic phototrophic bacterium Chloroflexus aurantiacus. Surprisingly, Mg-protoporphyrin IX monomethyl ester (oxidative) cyclase, AcsF, was identified under anaerobic growth conditions. The AcsF protein was found in the isolated chlorosome fractions, and the proteomics analysis suggested that significant portions of the AcsF proteins are not accessible to protease digestion. Additionally, quantitative real-time PCR studies showed that the transcript level of the acsF gene is not lower in anaerobic growth than in semiaerobic growth. Since the proposed enzymatic activity of AcsF requires molecular oxygen, our studies suggest that the roles of AcsF in C. aurantiacus need to be investigated further.

Robustness of electronic coherence in the Fenna–Matthews–Olson complex to vibronic and structural modifications

We present the first two-dimensional electronic spectra of photosynthetic antenna complexes bearing modifications to the protein and the chromophores. The vibronic structure of the Fenna-Matthews-Olson complex was altered by near-complete substitution of 13C for naturally abundant carbon and separately by randomly distributed partial deuteration. The structure and arrangement of the bacteriochlorophyll a chromophores were modified by deletion of the gene encoding the enzyme responsible for reducing the isoprenoid tail of the bacteriochlorophylls. Analysis of the time-dependent amplitude of the crosspeak corresponding to excitons 1 and 2 indicates that these modifications do not affect the frequency or dephasing of the beating observed in this particular peak. This result leads us to conclude that this beating indeed arises from electronic coherence and not vibrational wavepacket motion. We further conclude that the protection of zero-quantum coherences afforded by the protein matrix of this photosynthetic complex is not the result of a finely-tuned series of system-bath interactions perfected by billions of years of evolution but rather a simple downstream property of a close arrangement of chromophores within a phonon bath. We conclude with a brief discussion of the outstanding questions and possible applications of this phenomenon.

STRUCTURAL ANALYSIS OF ALTERNATIVE COMPLEX III IN THE PHOTOSYNTHETIC ELECTRON TRANSFER CHAIN OF CHLOROFLEXUS AURANTIACUS

The green photosynthetic bacterium Chloroflexus aurantiacus, which belongs to the phylum of filamentous anoxygenic phototrophs, does not contain a cytochrome bc or bf type complex which is found in all other known groups of phototrophs. This suggests that a functional replacement exists to link the reaction center photochemistry to cyclic electron transfer as well as respiration. Earlier work identified a potential substitute of the cytochrome bc complex, now named alternative complex III (ACIII), which has been purified from C. aurantiacus, identified, and characterized. ACIII functions as a menaquinol:auracyanin oxidoreductase in the photosynthetic electron transfer chain, and a related but distinct complex functions in respiratory electron flow to a terminal oxidase. In this work, we focus on elucidating the structure of photosynthetic ACIII. We found that ACIII is an integral membrane protein complex of approximately 300 kDa that consists of eight subunits of seven different types. Among them, there are four metalloprotein subunits, including a 113 kDa iron-sulfur cluster-containing polypeptide, a 25 kDa penta-heme c-containing subunit, and two 20 kDa monoheme c-containing subunits in the form of a homodimer. A variety of analytical techniques were employed in determining the ACIII substructure, including HPLC combined with ESI-MS, metal analysis, potentiometric titration, and intensity analysis of heme staining SDS-PAGE. A preliminary structural model of ACIII is proposed on the basis of the analytical data and chemical cross-linking in tandem with mass analysis using MALDI-TOF, as well as transmembrane and transit peptide analysis.