Function of the IsiA pigment–protein complex in vivo

Photosynthesis Research - The acclimation of cyanobacterial photosynthetic apparatus to iron deficiency is crucial for their performance under limiting conditions. In many cyanobacterial species,...     

PCS-based structure determination of protein–protein complexes

A simple and fast nuclear magnetic resonance method for docking proteins using pseudo-contact shift (PCS) and ¹H^N/¹⁵N chemical shift perturbation is presented. PCS is induced by a paramagnetic lanthanide ion that is attached to a target protein using a lanthanide binding peptide tag anchored at two points. PCS provides long-range (~40 Å) distance and angular restraints between the lanthanide ion and the observed nuclei, while the ¹H^N/¹⁵N chemical shift perturbation data provide loose contact-surface information. The usefulness of this method was demonstrated through the structure determination of the p62 PB1-PB1 complex, which forms a front-to-back 20 kDa homo-oligomer. As p62 PB1 does not intrinsically bind metal ions, the lanthanide binding peptide tag was attached to one subunit of the dimer at two anchoring points. Each monomer was treated as a rigid body and was docked based on the backbone PCS and backbone chemical shift perturbation data. Unlike NOE-based structural determination, this method only requires resonance assignments of the backbone ¹H^N/¹⁵N signals and the PCS data obtained from several sets of two-dimensional ¹⁵N-heteronuclear single quantum coherence spectra, thus facilitating rapid structure determination of the protein–protein complex.     

Low-temperature single-molecule spectroscopy on photosynthetic pigment–protein complexes from purple bacteria

The primary reactions of purple bacterial photosynthesis take place within two well characterized pigment–protein complexes, the core Reaction Center–Light Harvesting 1 (RC–LH1) complex and the more peripheral Light Harvesting 2 (LH2) complex. These antenna complexes serve to absorb incident solar radiation and to transfer it to the reaction-centers, where it is used to ‘power’ the photosynthetic redox reaction. This review provides an overview of how the character of the electronically excited states of these pigment–protein complexes are determined by quantum mechanics and how the respective spectral signatures can be observed by single-molecule spectroscopy.     

Interaction of pigment—protein complexes within aggregates stimulates dissipation of excess energy

Pigment—protein complexes in photosynthetic membranes exist mainly as aggregates that are functionally active as monomers but more stable due to their ability to dissipate excess energy. Dissipation of energy in the photosystem I (PSI) trimers of cyanobacteria takes place with a contribution of the long-wavelength chlorophylls whose excited state is quenched by cation radical of P700 or P700 in its triplet state. If P700 in one of the monomer complexes within a PSI trimer is oxidized, energy migration from antenna of other monomer complexes to cation radical of P700 via peripherally localized long-wave-length chlorophylls results in energy dissipation, thus protecting PSI complex of cyanobacteria against photodestruction. It is suggested that dissipation of excess absorbed energy in aggregates of the light-harvesting complex LHCII of higher plants takes place with a contribution of peripherally located chlorophylls and carotenoids.Translated from Biokhimiya, Vol. 69, No. 11, 2004, pp. 1592–1599.Original Russian Text Copyright © 2004 by Karapetyan     

Effects of Glycerol on the Fluorescence Spectra and Chloroplast Ultrastructure of Phaeodactylum tricornutum （Bacillariophyta）

Responses of the photosynthetic activity of Phaeodactylum tricornutum （Bacillariophyta） to organic carbon glycerol were investigated. The growth rate, photosynthetic pigments, 77 K fluorescence spectra, and chloroplast ultrastructure of P. tricornutum were examined under photoautotrophic, mixotrophic, and photoheterotrophic conditions. The results showed that the specific growth rate was the fastest under mixotrophic conditions. The cell photosynthetic pigment content and values of Chl a/Chl c were reduced under mixotrophic and photoheterotrophic conditions. The value of carotenoid/Chl a was enhanced under mixotrophic conditions, but was decreased under photoheterotrophic conditions. In comparison with photoautotrophic conditions, the fluorescence emission peaks and fluorescence excitation peaks were not shifted. The relative fluorescence of photosystem （PS） Ⅰ and PS Ⅱ and the values of F685/F710 and F685/F738 were decreased. Chloroplast thylakoid pairs were less packed under mixotrophic and photoheterotrophic conditions. There was a strong correlation between degree of chloroplast thylakoid packing and the excitation energy kept in PS Ⅱ. These results suggested that the PS Ⅱ activity was reduced by glycerol under mixotrophic conditions, thereby leading to repression of the photosynthetic activity.     

A model of the protein–pigment baseplate complex in chlorosomes of photosynthetic green bacteria

In contrast to photosynthetic reaction centers, which share the same structural architecture, more variety is found in the light-harvesting antenna systems of phototrophic organisms. The largest antenna system described, so far, is the chlorosome found in anoxygenic green bacteria, as well as in a recently discovered aerobic phototroph. Chlorosomes are the only antenna system, in which the major light-harvesting pigments are organized in self-assembled supramolecular aggregates rather than on protein scaffolds. This unique feature is believed to explain why some green bacteria are able to carry out photosynthesis at very low light intensities. Encasing the chlorosome pigments is a protein-lipid monolayer including an additional antenna complex: the baseplate, a two-dimensional paracrystalline structure containing the chlorosome protein CsmA and bacteriochlorophyll a (BChl a). In this article, we review current knowledge of the baseplate antenna complex, which physically and functionally connects the chlorosome pigments to the reaction centers via the Fenna–Matthews–Olson protein, with special emphasis on the well-studied green sulfur bacterium Chlorobaculum tepidum (previously Chlorobium tepidum). A possible role for the baseplate in the biogenesis of chlorosomes is discussed. In the final part, we present a structural model of the baseplate through combination of a recent NMR structure of CsmA and simulation of circular dichroism and optical spectra for the CsmA–BChl a complex.     

Modeling glycosaminoglycan–protein complexes

Glycosaminoglycans are long linear and complex polysaccharides that are fundamental components of the mammalian extracellular matrix. Therefore, it is crucial to appropriately characterize molecular structure, dynamics, and interactions of protein-glycosaminoglycans complexes for improving understanding of molecular mechanisms underlying GAG biological function. Nevertheless, this proved challenging experimentally, and theoretical techniques are beneficial to construct new hypotheses and aid the interpretation of experimental data. The scope of this mini-review is to summarize four specific aspects of the current theoretical approaches for investigating noncovalent protein-glycosaminoglycan complexes such as molecular docking, free binding energy calculations, modeling ion impact, and addressing the phenomena of multipose binding of glycosaminoglycans to proteins.     

Effect of protein aggregation on the spectroscopic properties and excited state kinetics of the LHCII pigment–protein complex from green plants

Steady-state and time-resolved absorption and fluorescence spectroscopic experiments have been carried out at room and cryogenic temperatures on aggregated and unaggregated monomeric and trimeric LHCII complexes isolated from spinach chloroplasts. Protein aggregation has been hypothesized to be one of the mechanistic factors controlling the dissipation of excess photo-excited state energy of chlorophyll during the process known as nonphotochemical quenching. The data obtained from the present experiments reveal the role of protein aggregation on the spectroscopic properties and dynamics of energy transfer and excited state deactivation of the protein-bound chlorophyll and carotenoid pigments.     

Near edge X-ray absorption fine structure spectroscopy (NEXAFS) of pigment–protein complexes: Peridinin–chlorophyll a protein (PCP) of Amphidinium carterae

Peridinin–chlorophyll a protein (PCP) is a unique water soluble antenna complex that employs the carotenoid peridinin as the main light-harvesting pigment. In the present study the near edge X-ray absorption fine structure (NEXAFS) spectrum of PCP was recorded at the carbon K-edge. Additionally, the NEXAFS spectra of the constituent pigments, chlorophyll a and peridinin, were measured. The energies of the lowest unoccupied molecular levels of these pigments appearing in the carbon NEXAFS spectrum were resolved. Individual contributions of the pigments and the protein to the measured NEXAFS spectrum of PCP were determined using a “building block” approach combining NEXAFS spectra of the pigments and the amino acids constituting the PCP apoprotein. The results suggest that absorption changes of the pigments in the carbon near K-edge region can be resolved following excitation using a suitable visible pump laser pulse. Consequently, it may be possible to study excitation energy transfer processes involving “optically dark” states of carotenoids in pigment–protein complexes by soft X-ray probe optical pump double resonance spectroscopy (XODR).     

Analysis of secondary structural and physicochemical changes in protein–protein complexes

Conformation switching in protein–protein complexes is considered important for the molecular recognition process. Overall analysis of 123 protein–protein complexes in a benchmark data-set showed that 6.8% of residues switched over their secondary structure conformation upon complex formation. Amino acid residue-wise preference for conformation change has been analyzed in binding and non-binding site residues separately. In this analysis, residues such as Ser, Leu, Glu, and Lys had higher frequency of secondary structural conformation change. The change of helix to coil and sheet to coil conformation and vice versa has been observed frequently, whereas the conformation change of helix to extended sheet occurred rarely in the studied complexes. Influence of conformation change toward the N and C terminal on either side of the binding site residues has been analyzed. Further, analysis on φ and ψ angle variation, conservation, stability, and solvent accessibility have been performed on binding site residues. Knowledge obtained from the present study could be effectively employed in the protein–protein modeling and docking studies.     

Purification and characterization of HIV–human protein complexes

To fully understand how pathogens infect their host and hijack key biological processes, systematic mapping of intra-pathogenic and pathogen–host protein–protein interactions (PPIs) is crucial. Due to the relatively small size of viral genomes (usually around 10–100 proteins), generation of comprehensive host–virus PPI maps using different experimental platforms, including affinity tag purification-mass spectrometry (AP-MS) and yeast two-hybrid (Y2H) approaches, can be achieved. Global maps such as these provide unbiased insight into the molecular mechanisms of viral entry, replication and assembly. However, to date, only two-hybrid methodology has been used in a systematic fashion to characterize viral–host protein–protein interactions, although a deluge of data exists in databases that manually curate from the literature individual host–pathogen PPIs. We will summarize this work and also describe an AP-MS platform that can be used to characterize viral-human protein complexes and discuss its application for the HIV genome.     

Modeling of the structure of protein–DNA complexes using the data from FRET and footprinting experiments

We discuss the question of constructing three-dimensional models of DNA in complex with proteins using computer modeling and indirect methods of studying the conformation of macromolecules. We consider the methods of interpreting the experimental data obtained by indirect methods of studying the three-dimensional structure of biomolecules. We discuss some aspects of integrating such data into the process of constructing the molecular models of DNA–protein complexes based on the geometric characteristics of DNA. We propose an algorithm for estimating conformations of such complexes based on the information about the local flexibility of DNA and on the experimental data obtained by Forster resonance energy transfer (FRET) and hydroxyl footprinting. Finally, we use this algorithm to predict the hypothetical configuration of DNA in a nucleosome bound with histone H1.     

Computation of temperature-dependent dissociation rates of metastable protein–ligand complexes

Molecular simulations are often used to analyse the stability of protein–ligand complexes. The stability can be characterised by exit rates or using the exit time approach, i.e. by computing the expected holding time of the complex before its dissociation. However determining exit rates by straightforward molecular dynamics methods can be challenging for stochastic processes in which the exit event occurs very rarely. Finding a low variance procedure for collecting rare event statistics is still an open problem. In this work we discuss a novel method for computing exit rates which uses results of Robust Perron Cluster Analysis (PCCA+). This clustering method gives the possibility to define a fuzzy set by a membership function, which provides additional information of the kind ‘the process is being about to leave the set’. Thus, the derived approach is not based on the exit event occurrence and, therefore, is also applicable in case of rare events. The novel method can be used to analyse the temperature effect of protein–ligand systems through the differences in exit rates, and, thus, open up new drug design strategies and therapeutic applications.     

The fluorescence yield of the trimeric fucoxanthin–chlorophyll–protein FCPa in the diatom Cyclotella meneghiniana is dependent on the amount of bound diatoxanthin

The fluorescence yield of isolated fucoxanthin chlorophyll proteins, serving as light harvesting proteins in diatoms, was compared to the amount of diatoxanthin bound. Diatoxanthin was earlier shown to be involved in the xanthophyll cycle in diatoms as a functional analogue of zeaxanthin in higher plants. By growing cells under different light conditions, the amount of diatoxanthin in both the trimeric FCPa as well as the oligomeric FCPb of the diatom Cyclotella meneghiniana was increased. In the trimeric FCPa, the fluorescence yield decreased with increasing diatoxanthin content, whereas in the oligomeric FCPb fluorescence was generally lower, albeit constant. No pH dependence of fluorescence yield could be demonstrated except for artificially aggregated FCPa. Thus, diatoxanthin is able to quench fluorescence in FCPa, but the yield is also influenced by pH when the protein becomes aggregated.