Fluorescence bacteriochlorophyll

Bacteriochlorophyll in Chromatium has three absorption bands with peak positions at 800, 850, and 890 nm. The last includes the reaction center bacteriochlorophyll and is the only form that fluoresces. Recent studies have established that most if not all chlorophyll is bound to specific proteins, a fact that can account for the various overlapping absorption bands. 

Figure 23-26 (A), (B) Arrangement of bacteriochlorophyll chromophores in the cyclic LH2 array of Rhodopseudomonas acidophila. The B850 subunits are gray while the B800 subunits are black. (C) Fluorescence-excitation spectra. Top trace, for an ensemble of LH2 complexes, other traces, for several individual LH2 complexes at 1.2K. Fine structure is evident for the B800 but not for the B850 chromophores. From van Oijen et al.299 with permission.

Reaction center kinetics. After an 0.8-ps or shorter flash of light the decay of the singlet excited state of the bacteriochlorophyll dimer in isolated reaction centers can be followed by loss of its characteristic fluorescence.328 329 The lifetime of this excited state in R. sphaeroides is only 4 ps indicating a rapid occurrence of the initial electron transfer of Eq. 23-31. 

Table 2 shows the location of the carotenoid band in the spectra (absorption and excitation of bacteriochlorophyll fluorescence) of R... 

TABLE 2 Correspondence between the carotenoid bands in the absorption and the bacteriochlorophyll-fluorescence excitation spectra of R rubrum cells... 

Very fast electron transfers from P+ to bacteriochlorophyl (Bchl) and from (Bchl)- to QA do not depend on media dynamics and occur via conformationally non-equilibrium states (Fig.3.18). The dual fluorophore-nitroxide molecules (D-A) are also convenient objects for analysing the activity-dynamics relationship. The marked irreversible photoreduction of the nitroxide fragment of the dual probe incorporated into the binding site of HSA only took place when the nanosecond dynamical processes around the probe traced by ESR and fluorescence methods were detected (Rubtsova et al., 1993, Fogel et al, 1994 Likhtenshtein, 1986 Lozinsky et al., 2002). Similar results were reported for another model protein system, i.e. a-chymotrypsin with spin labeled methionin-92 groups (Belonogova et al., 1997). In the latter enzyme, the excited tryptophan group serves as an electron donor. 

The fluorescence emission of chlorophylls and bacteriochlorophylls is invariably emitted from the lowest energy excited sate, with typical Stokes shifts of 10-15 run from the Qy absorbance maximum. Fluorescence lifetimes of 2-5 ns are usually observed for monomeric pigments in organic solvents. Fluorescence of chlorophyll and bacteriochlorophyll in vivo is very strongly quenched excited-state lifetimes are typically a few tens of picoseconds and fluorescence quantum yields are at most a few percent. The... 

In a book written by specialists in the various areas of photosynthesis research, there are bound to be some overlaps and some gaps. One area that may not have been adequately covered, althovigh its impact can be discerned in various chapters, is the wealth of information regarding energy and electron transfer and structure derived from studies of prompt and delayed fluorescence of chlorophyll and bacteriochlorophyll. However, the reader interested in this area should find enough information in this book for further literature on the subject. 

Theoretical calculations have been made for photosynthetic pigments . An extensive review of models of energy and electron transfer events of synthetic molecules for photosynthesis has been prepared by Wasielewski . Other studies have made on tetraphenylporphyrin-polypeptide pigments , photosensitization of triplet carotenoids , fluorescence yields and lifetimes for bacteriochlorophyll c , triplet yields and ESR of chlorophyll 3 8 and quenching processes of pheophytin 539 ... 

The spectrum of the light emitted by tungsten / halogen lamps is better than the one from fluorescent lamps, taking into account that phototrophic bacteria use infrared light, with absorption maxima at 800 and 850 nm corresponding to the absorption of bacteriochlorophylls. 

Fig. 2. Model forthe chlorosome in Chloroflexus aurantiacus. Model adapted from Mimuro, Hirota, Nishimura, Moriyama, Yamazaki, Shimada, and Matsuura (1994) Molecular organization of bacteriochlorophylls in chlorosomes of the green photosynthetic bacteria Chloroflexus aurantiacus Studies of fluorescence depolarization accompanied with the energy transfer process. Photosynthesis Res. 41 190.

Bacteriochlorophyll c extracted from green bacteria has indeed been found to form oligomers in nonpolar solvents with absorption and fluorescence spectra matching those of chlorosomes in vivo. A BChl-c oligomer was specifically suggested as a component in a model for a chlorosome. 

The third example is the light-harvesting complex B800-850 obtained from the wild-type Rb. sphaeroides 2.4.1. This complex contains three BChls and one carotenoid (spheroidene) per protein subunit. The fluorescence excitation was obtained by monitoring the emission at 850 nm. Fig. 3 (C) shows the absorption and fluorescence excitation spectra in the 400-620 nm region, with the two spectra normalized at 590 nm (marked with ). The excellent match ofthe absorption and excitation spectra indicates that photoexcited spheroidene transfers energy to bacteriochlorophyll with a high efficiency. 

Clearly, this spectroscopic phenomenon was a serendipitous event because there was no corresponding selection applied during the generation of FluA. Also, the other mutants that were selected along with it did not show fluorescence quenching to the same high extent. Nevertheless, it is remarkable that such an efficient electron transfer process, which is even faster (by a factor 3-4) than the one measured between bacteriochlorophyll and bacteriopheophytin in the bacterial reac-... 

Frank HA, McGann WJ, Machniki J and Felber M (1982b) Magnetic field effects on the fluorescence of two reaction centerless mutants of Rhodopseudomonas capsulata. Biochem Biophys Res Comm 106 1310-1317 Frank HA, Machnicki J and Friesner R (1983) Energy transfer between the primary donor bacteriochlorophyll and carotenoids in Rhodopseudomonas sphaeroides. Photochem Photobiol 38 451-455... 

An attempt to produce films of chromatophores was made by Clayton by drying aqueous suspensions onto solid supports. Dehydration of RC from Rhodopseudomonas sphaeroides alters their adsorption spectra, photochemical efficiency, fluorescence yield and reaction kinetics of electrons that return from reduced ubiquinone to oxidized bacteriochlorophyll. The properties of the air-dried films were restored by exposure to water vapour. It suggests that water helps to preserve an optimal configuration for photochemical efficiency. 

Recently it was found that similar subunit forms could be obtained from the core antenna of both Rhodobacter sphaeroides and Rhodopseudomonas capsulata []. The B820 subunit form thus seems to be a basic form present in at least three different core antennae. This emphasizes the importance of elucidating the specific conformation of this subunit. We used low temperature fluorescence polarization measurements in combination with other spectroscopic techniques (OD and CD) to obtain information on the specific organization of the bacteriochlorophyll pigments in the subunit forms. 

Teuchner K, Stiel H, Leupold D, Scherz A, Noy D, Simonin I, Hartwich G, and Scheer H. Fluorescence and excited state absorption in modified pigments of bacterial photosynthesis - A comparative study of metal-substituted bacteriochlorophylls a. J. Luminesc. 1997 72-4 612-614. 

Wang, R.T., Clayton, R.K. Absolute yield of bacteriochlorophyll fluorescence in vivo. Photochem. Photobiol. 13, 215-224 (1971)... 

These expressions come into play if the molecule relaxes to a different excited electronic state before it fluoresces. In bacteriochlorophyll, for example, the transition dipole for excitation to the second excited state (Q ) is perpendicular to the transition dipole for the lowest state (Q, ). If a solution of bacteriochlorophyll in a viscous solvent such as glycerol is excited in the band at 575 nm, the emission from the band near 800 nm has an anisotropy of -0.2, which is the result given by Eq. (5.68) for = 90°. 

For an angle of 45°, Eq. (5.68) gives E = 0.1. An angle of 45° imphes that the emission transition dipole, on average, has equal projections on the absorptiOTi transition dipole and on another axis that is orthogonal to the absorptimi dipole. An equivalent situation can arise if the excited molecule rapidly transfers its energy to an array of other molecules whose transition dipoles lie in a plane but take oti all orientations within that plane. Some photosynthetic bacterial antenna systems contain such arrays of bacteriochlorophyll molecules. Following equilibration of the excitation over the array, fluorescence occurs with an anisotropy of 0.1 [175-177]. WeTl return to the fluorescence anisotropy of such multimolecular systems in Chap. 10. 

---