Structure, Function and Reconstitution of Antenna Complexes of Green Photosynthetic Bacteria

Most chlorophyll-type pigments in a photosynthetic organism function as an antenna, absorbing light and transferring excitations to a photochemical reaction center where energy storage takes place by a series of chemical reactions. The green photosynthetic bacteria are characterized by large antenna complexes known as chlorosomes (Fig. 2), in which pigment-pigment interactions are of dominant importance (4). The overall objective of this project is to determine structural data on the chlorosome, along with mechanisms of excitation transfer and regulation of this unique antenna system, including how it is integrated into the rest of the photosynthetic energy transduction apparatus. Techniques that have been used in this research include biochemical analysis, spectroscopy, several types of electron microscopy, X-ray diffraction, and reconstitution from purified components. Recent results indicate that the chlorosome baseplate structure, which is the membrane attachment site for the chlorosome to the membrane, is a unique pigment-protein that contains large amounts of carotenoids and small amounts of bacteriochlorophyll a. Reconstitution of directed energy transfer in chlorosomes is being investigated using purified baseplates and oligomeric pigments. The integral membrane B808-866 antenna complex from Chloroflexus aurantiacus and the Fenna-Matthews-Olson proteinreaction center complex from green sulfur bactreria are being characterized by spectroscopic and structural techniques.

Fig. 2 Three-dimensional electron microscopic tomography model of green sulfur bacteria.

First reported by our group, the auracyanins represent a novel class of small blue copper proteins (5). They are found in the photosynthetic bacterium Chloroflexus aurantiacus, which produces two distinct but closely related forms: Auracyanin A and AuracyaninB. Based on their redox potentials, their similarity to other cupredoxins, and the lack of any iron-based soluble electron carriersin C. aurantiacus, auracyanins most likely function as electron carriers in the periplasmic electron-transfer step of photosynthesis.

Fig. 3 Structure of auracyanin B.Recently, two cytochrome-containing complexes from C. aurantiacus were found to be members of a new class of bacterial membrane oxidoreductases, and are candidates for electron donors to auracyanins (see next section). The functional significance of two forms of auracyanins in C. aurantiacus is currently under investigation. A 1.55Å crystal structure of auracyanin B has been reported (5), and the structure of auracyanin A is also now complete (6). The genes for both proteins have been cloned in bacterial expression systems allowing overexpression of large quantities of proteins for further biochemical studies. We measure protein expression levels using western blots employing antibodies specific for either auracyanin A or B, as well as quantitative real-time RT-PCR of auracyanin mRNAs. We are also studying the copper active sites using UV-Vis, electron paramagnetic and X-ray absorption spectroscopy. A project that involves site-directed mutations in both Aura A and Aura B is planned, and will focus on the relationship of structure and spectroscopic signatures.