Neil W. Isaacs

The Structure and Thermal Motion of the B800–850 LH2 Complex from Rps. acidophila at 2.0 Å Resolution and 100 K: New Structural Features and Functionally Relevant Motions

The structure at 100K of integral membrane light-harvesting complex II (LH2) from Rhodopseudomonas acidophila strain 10050 has been refined to 2.0A resolution. The electron density has been significantly improved, compared to the 2.5A resolution map, by high resolution data, cryo-cooling and translation, libration, screw (TLS) refinement. The electron density reveals a second carotenoid molecule, the last five C-terminal residues of the alpha-chain and a carboxy modified alpha-Met1 which forms the ligand of the B800 bacteriochlorophyll. TLS refinement has enabled the characterisation of displacements between molecules in the complex. B850 bacteriochlorophyll molecules are arranged in a ring of 18 pigments composed of nine approximate dimers. These pigments are strongly coupled and at their equilibrium positions the excited state dipole interaction energies, within and between dimers, are approximately 370cm(-1) and 280cm(-1), respectively. This difference in coupling energy is similar in magnitude to changes in interaction energies arising from the pigment displacements described by TLS tensors. The displacements appear to be non-random in nature and appear to be designed to optimise the modulation of pigment energy interactions. This is the first time that LH2 pigment displacements have been quantified experimentally. The calculated energy changes indicate that there may be significant contributions to inter-pigment energy interactions from molecular displacements and these may be of importance to photosynthetic energy transfer.

Structure‐Based Calculations of the Optical Spectra of the LH2 Bacteriochlorophyll‐Protein Complex from Rhodopseudomonas acidophila

Abstract— The molecular structure of the light‐harvesting complex 2 (LH2) bacteriochlorophyll‐protein antenna complex from the purple non‐sulfur photosynthetic bacterium Rhodopseudomonas acidophila, strain 10050 provides the positions and orientations of the 27 bacteriochlorophyll (BChl) molecules in the complex. Our structure‐based model calculations of the distinctive optical properties (absorption, CD, polarization) of LH2 in the near‐infrared region use a point‐monopole approximation to represent the BChl Qy transition moment. The results of the calculations support the assignment of the ring of 18 closely coupled BChl to B850 (BChl absorbing at 850 nm) and the larger diameter, parallel ring of 9 weakly coupled BChl to B800. All of the significantly allowed transitions in the near infrared are calculated to be perpendicular to the C9 symmetry axis, in agreement with polarization studies of this membrane‐associated complex. To match the absorption maxima of the B800 and B850 components using a relative permittivity (dielectric constant) of 2.1, we assign different site energies (12 500 and 12260 cm−1, respectively) for the Qy transitions of the respective BChl in their protein binding sites. Excitonic coupling is particularly strong among the set of B850 chromophores, with pairwise interaction energies nearly 300 cm between nearest neighbors, comparable with the experimental absorption bandwidths at room temperature. These strong interactions, for the full set of 18 B850 chromophores, result in an excitonic manifold that is 1200 cm−1 wide. Some of the upper excitonic states should result in weak absorption and perhaps stronger CD features. These predictions from the calculations await experimental verification.

The two types of 3-dehydroquinase have distinct structures but catalyze the same overall reaction.

The structures of enzymes catalyzing the reactions in central metabolic pathways are generally well conserved as are their catalytic mechanisms. The two types of 3-dehydroquinate dehydratase (DHQase) are therefore most unusual since they are unrelated at the sequence level and they utilize completely different mechanisms to catalyze the same overall reaction. The type I enzymes catalyze a cis-dehydration of 3-dehydroquinate via a covalent imine intermediate, while the type II enzymes catalyze a trans-dehydration via an enolate intermediate. Here we report the three-dimensional structures of a representative member of each type of biosynthetic DHQase. Both enzymes function as part of the shikimate pathway, which is essential in microorganisms and plants for the biosynthesis of aromatic compounds including folate, ubiquinone and the aromatic amino acids. An explanation for the presence of two different enzymes catalyzing the same reaction is presented. The absence of the shikimate pathway in animals makes it an attractive target for antimicrobial agents. The availability of these two structures opens the way for the design of highly specific enzyme inhibitors with potential importance as selective therapeutic agents.

A model for the photosynthetic apparatus of purple bacteria

Abstract The photosynthetic apparatus of purple bacteria is composed of light-harvesting complexes and reaction centres. Recent work on the structures of light-harvesting complexes combined with existing knowledge of the structure of the reaction centre now makes it feasible, for the first time, to model the entire photosynthetic apparatus. Questions can be asked about the functional implications of such a model in the light of the most recent spectroscopic data.

The purple bacterial photosynthetic unit

Now is a very exciting time for researchers in the area of the primary reactions of purple bacterial photosynthesis. Detailed structural information is now available for not only the reaction center (Lancaster et al. 1995, in: Blankenship RE et al. (eds) Anoxygenic Photosynthetic Bacteria, pp 503–526), but also LH2 from Rhodopseudomonas acidophila (McDermott et al. 1995, Nature 374: 517–521) and LH1 from Rhodospirillum rubrum (Karrasch et al. 1995. EMBO J 14: 631–638). These structures can now be integrated to produce models of the complete photosynthetic unit (PSU) (Papiz et al., 1996, Trends Plant Sci, in press), which opens the door to a much more detailed understanding of the energy transfer events occurring within the PSU.

Probing the interface between membrane proteins and membrane lipids by X-ray crystallography

Biological membranes are composed of a complex mixture of lipids and proteins, and the membrane lipids support several key biophysical functions, in addition to their obvious structural role. Recent results from X-ray crystallography are shedding new light on the precise molecular details of the protein-lipid interface.

Rings, ellipses and horseshoes: how purple bacteria harvest solar energy.

This Review summarises the current state of research on the structure and function of light-harvesting apparatus in purple photosynthetic bacteria. Particular emphasis is placed on the major open questions still outstanding in this field in addition to what is already known.

Purification and biochemical characterization of the D6 chemokine receptor.

There is much interest in chemokine receptors as therapeutic targets in diseases such as AIDS, autoimmune and inflammatory disorders, and cancer. Hampering such studies is the lack of accurate three-dimensional structural models of these molecules. The CC-chemokine receptor D6 is expressed at exceptionally high levels in heterologous transfectants. Here we report the purification and biochemical characterization of milligram quantities of D6 protein from relatively small cultures of transfected mammalian cells. Importantly, purified D6 retains full functional activity, shown by displaceable binding of 125I-labelled MIP-1beta (macrophage inflammatory protein-1beta) and by complete binding of the receptor to a MIP-1alpha affinity column. In addition, we show that D6 is decorated on the N-terminus by N-linked glycosylation. Mutational analysis reveals that this glycosylation is dispensable for ligand binding and high expression in transfected cells. Metabolic labelling has revealed the receptor to also be sulphated and phosphorylated. Phosphorylation is ligand independent and is not enhanced by ligand binding and internalization, suggesting similarities with the viral chemokine receptor homologue US28. Like US28, an analysis of the full cellular complement of D6 in transfected cells indicates that >80% is found associated with intracellular vesicular structures. This may account for the high quantities of D6 that can be synthesized in these cells. These unusual properties of D6, and the biochemical characterization described here, leads the way towards work aimed at generating the three-dimensional structure of this seven-transmembrane-spanning receptor.

Creating permanent 3D arrangements of isolated cells using holographic optical tweezers

We report the creation of permanent 3D configurations of cells, at predefined positions, within a gelatin matrix. The technique used holographic optical tweezers to manipulate individual E. coli within a solution comprising monomer precursors. The matrix was then set and after the laser beam was removed, we were able to demonstrate that the structures remained intact for many days. We were also able to demonstrate that, in the presence of appropriate nutrients, the E. coli survived within the gelatin matrix for several days. The technique could have a number of potential future applications, including the arrangement of a variety of different cell types in complex architectures, as motifs for promoting tissue differentiation and growth within the field of cell engineering.

Structural factors which control the position of the Qy absorption band of bacteriochlorophyll a in purple bacterial antenna complexes

This paper presents a concise review of the structural factors which control the energy of the Qy absorption band of bacteriochlorophyll a in purple bacterial antenna complexes. The energy of these Qy absorption bands is important for excitation energy transfer within the bacterial photosynthetic unit.