Bacterial Photosynthesis

Photocheinistryand Photobiology Vol. 38. No. 6, pp. 769-112.1983 Printed in Grcat Britain. All rights reserved

003 I -X655183$(~3.(N)+O.(K) Copyright 01983 Pergamon PressLtd

YEARLY REVIEW

BACTERIAL PHOTOSYNTHESIS INTRODUCTION

The light reactions of bacterial photosynthesis (that is from the initial absorption of a photon right on through to the generation of ATP and reduced pyridine nucleotides) have been subjected to extensive biophysical analyses (see, for example, Clayton and Sistrom, 1978). Unfortunately, in numerous cases, unambiguous interpretation of these data has been prevented by a lack of fundamental biochemical information. What has been missing is either detail on the composition or the structure of the various preparations under study, or indeed, both. However, in the last year or so, there have been several significant biochemical advances that now promise to rectify this situation. It is these areas of bacterial photosynthesis which will be considered in this Yearly Review.

Reaction centres The primary photochemical reaction in bacterial photosynthesis has recently been expertly reviewed by Parson (1982). Bacterial reaction centres are well defined pigment-protein complexes (except from green sulphur photosynthesic bacteria where no reaction centre is yet available) and usually consist of four molecules of bacteriochlorophyll, two molecules of bacteriopheophytin, one molecule of carotenoid, one or two molecules of quinone and one atom of ferrous iron, all bound by three polypeptides (called ‘H’, ‘M’ and ‘L’). T h e reaction centre from Cliloroflexus aurantiacus is an exception to this general picture (Pierson and Thornber, 1983; Pierson et al., 1983). Largely because the composition of the bacterial reaction centres is so well characterized, the kinetics of the primary reaction are well understood. But even in this case, a lack of structural information on the spacial arrangement of the reaction centre pigments has been a major block to further progress. It is, however, in this very area that perhaps this year’s most exciting development in research into bacterial photosynthesis has occurred. Michel(l982) has reported the crystallisation of reaction centres from Rhodopseudomonas viridis. The crystals are large tetragonal ones, have the space group P4,, 2 , , 2 and probably contain a single reaction centre per unit cell (Zinth etal., 1983). They have a delightful brown colour and defract X-rays clearly to beyond 2.5 A. Heavy atom derivatives are now being collected (H. Michel, personal communication) and a successful structural determination is impatiently awaited. It will truly b e a l a n d m a r k in t h e s t u d y of

photosynthesis if the structure determined by X-ray crystallography is able t o reveal the precise 3-dimensional arrangement of all the constituent reaction centre pigment molecules. It may, however, be much harder to assign which pigments in the structure correspond to which of the reaction centre absorption bands. So far, the prospects seem favourable, but, it should be pointed out that in crystallographic terms Rps. viridis reaction centres are rather large. They contain four polypeptides, the well-known ‘H’, ‘M’ and ‘L’ subunits and a c-type cytochrome (s) (Thronber etal. ,1980). This probably m e a n s t h a t t h e primary s t r u c t u r e of these p o l y p e p t i d e s will b e r e q u i r e d f o r a full high-resolution structure. Although reaction centres from Rps. viridis contain bacteriochlorophyll b rather than the more usual bacteriochlorophyll a , it is extremely likely that information about the structure of Rps. viridis reaction centres will be of general significance since the pattern of their primary reactions are nearly identical to those in the bacteriochlorophyll a containing reaction centres (Parson, 1982). There have been several efforts made to try and determine the amino acid sequence of the reaction centre polypeptides using conventional protein sequencing techniques (Sutton et al., 1982; Theiler. 1982; and H. Michel, personal communication). So far these attempts have been time consuming and h a v e yielded only r a t h e r limited s e q u e n c e information on the first 3&50 amino acids at the amino termini of the ‘H’, ‘M’ and ‘L’ polypeptides. More recently this problem has been tackled utilising the more modern techniques of molecular biology. (Williams etal., 1983; Youvan et a / . , 1983). Williams er a l . ( 1 9 8 3 ) h a v e c l o n e d t h e g e n e f r o m Rhodopseudomonas shaeroides for the ‘M’ subunit of the reaction centre and have almost completed its sequence. In Rhodopseudomonas capsulata Youvan eral. (1983) have cloned the genes of the ‘H’, ‘M’ and ‘L’ subunits and have again almost completed their sequences. The results of these studies are also eagerly awaited. It is to be hoped that a similar approach will be successful with the reaction centre polypeptides of Rps. viridis, and will be in time to be of use in t h e i n t e r p r e t a t i o n of t h e X-ray crystallography data.

Light-harvesting pigment-protein complexes The structure and function of a range of bacterial light-harvesting complexes has recently been

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Yearly Review

reviewed by Thornber er ul. (1983). Interestingly, as in the case of research into reaction centres, this past year has seen an exciting new development in this area.Zuber's group in Zurich. by using an organic solvent extraction technique. first suggested by Tonn et ul. (1977). have sequenced a range of bacterial antenna polypeptides. So far the data is available on four polypeptides from Rps. capsulata (Tadros et a/. 1983: Theiler et 01.. 1982). four polypeptides from Rps. sphaeroides (Theiler et al., 1983), two p o I y p e p t i d t's f r o m R h o d o s p ir illu rn r u b r u m (Brunisholz e r al.. 1981; Theiler et al.. 1983). three polypeptides from Rps. riridis (Brunisholz et al., 1YX3a) and one from Rhodopseudomonas gelatinosa (Theiler et d . . 1982). A comparison of these hequences has led to a number of interesting predictions a b o u t t h e i r p r o b a b l e secondary 5tructure. a n d these s h o u l d b e helpful f o r interpreting future functional studies. So far every polypeptide has a conserved histidine residue in the centre of a hydrophobic region. It has been suggested t h a t t h i s m a y b e t h e fifth l i g a n d of t h e bacteriochlorophyll. All the polypeptides are small (3-7 k D ) and very hydrophobic (hence their solubility in organic solvents). Each type of antenna complex appears to contain a pair of non-identical pigment-binding polypeptides which are thought to form a fundamental 'minimal unit'. This unit then aggregates to form the larger. native in vivo complex (Cogdell and Thornber. 1980). Without exception, these polypeptides show three distinct structural domains. being polar at each end and non-polar in the centre. This has naturally led to the prediction that each polypeptide should lie across t h e photosynthetic membrane with the central region probably folded into an a helix (Zuber et al., 1983). Analysih of the B80&-850 antenna complex from Rps. sphaeroitles hy both CD in the far UV (Cogdell and Scheer. unpublished observations) and by attenuated total reflection I R spectroscopy (Theiler and Zuber, 1983) has indeed confirmed the presence of a large amount of u helical structure. There have been several recent studies designed to determine the membrane topology of the antenna complexes and the reaction centres (for example. Bachmann er ul.. 1YX1: Cuendet et al., 1978; Peters and Drews. 1983). The aim of these studies is to try and huild up a good model for the structure of the photosynthetic unit. This can then be used in the interpretation of studies designed to investigate the mechanims o f energy transfer within the in vivo pigment bed. With the availability of the sequence data the resolution that is possible with these studies is greatly increased. I t is now possible to exactly determine the \ites t > f cheniical modification or proteolytic attack. A p o d ruample of this is provided in the study of Brunisholz et rrl. ( 198%) who investigated the effect of Prutrinase K upon the B 890 light-harvesting polypeptides in R. rubrurn chromataphores.

Proteinase K removed the first six amino acids from the amino terminal of one polypeptide and the first sixteen from the amino terminal of the second polypeptide. In both cases the carboxyterminus was unaffected. This study reveals that not only are the amino termini of both antenna polypeptides present on the cytoplasmic surface of the membrane, but also illustrates the precision with which topological information can now be obtained.

The blcl complexes Following charge separation within t h c photochemical reaction centre, the electron on the p r i m a r y e l e c t r o n - a c c e p t o r e n t e r s a cyclic electron-transport pathway, eventually returning to the reaction centre and reducing the oxidised primary donor (Clayton and Sistrom, 1978). The core of this cyclic electron transport pathway is the b/cl complex (Hauska et al.. 1983). The kinetics of cyclic electron transport and the mechanism of its coupled energy conservation reactions have been the subject of extensive biophysical studies (for example, see Clayton and Sistrom. 1978). However, in all these studies there has been the underlying problem that the basic biochemical composition of the blcl region of the electron transport chain was very poorly defined. This area of photosynthesis research has been considerably clarified now that the bic, complex has been isolated and partially characterized from a photosynthetic bacterium Rps. sphaeroides strain G A (Gabellini et a / . , 1983a and b; 1982). It is now clear that there is a basic structural and functional similarity between the complexes isolated from mitochondria, chloroplasts and photosynthetic bacteria (Crofts et al., 1982; Hauska, 1983). The blc, complex from Rps. sphaeroides strain G A contains four major polypeptides (Gabellini, 1982): 40 kd (cytochrome b ) , 34 kd (cytochrome c , ) see also Wood (Wood, 1980). 25 kd (probably the Rieske FeSprotein) and one of 6 kd (function unassigned). T h e complex also contains some ubiquinone (Gabellini and Hauska 1983b). The cytochrome b has been purified and partially characterized (Gabellini and Hauska, 1983a and b ) . As would be expected, it shows many similarities to the cytochrome b found in the mitochondnal b / c l , complex, e.g. mid-point potential heterogeneity and p H dependence (Gabellini and Hauska, 1983a: von Jagow and Engel, 1981). As more information is being compiled about the various components of the b l c l , complex, the topology within the membrane is being studied. The Rieske iron-sulphur cluster is asymmetrically placed in the membrane near the cytochrome c1 side (Prince, 1983). Further work using antibodies to purified components and reconstruction experiments analogous to those carried out with mitochrondrial complexes (Engel et al., 1983; Mendel-Hartvig and

Yearly Review

Nelson. 1983) should provide further topological information. In view of the structural and functional similarities between the complexes found in mitochrondria, chloroplasts and bacteria it is now possible to collate the data collected from these complexes to aid the understanding of the bacterial photosynthetic cyclic electron transport pathway. A detailed description of the b/cl. complex is beyond the scope of this review, but the excellent paper by Hauskaetal. (1983) should be consulted by those interested in this area.

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11. Gabellini. N. and G . Hauska (1983a) Characterization of cytochrome b in the isolated ubiquinol- cytochrome c o xid o r e d u c t a s e f r o m R h o d o p s e u d om o n 0.r sphaeroides G A . FEBS Lett. 153, 146150. 12. Gabellini. N. and G . Hauska (1983b) Isolation of cytochrome b from the cytochrome bc, complex of Rhodopseudomonas sphaeroides GA. FEBS Let[. 154, 171-174. 13 Hauska, G., E. Hurt. N. Gabellini and W. Lockau (1983) Comparative aspects of quinol-cytochrom c/plastocyanin oxidoreductases. Biochirn. HiophyJ. Acra 726. 97-133. 14 Mendel-Hartvig. I. and B.D. Nelson (1983) Studics on beef heart ubiquinol-cytochrome c reductase. Topological studies on the core proteins using proteolytic RICHARD J. COGDELL digestion and immunreplication. J . Bioenerget. JANEVALENTINE Biomembr. 15, 27-36. 15 Michel, H . (1982) Three-dimensional crystals 01 I Department of Botany membrane protein complex: The photochemical reacUniversity of Glasgow tion centre from Rhodopseudomonas viridis. J . Mol. Glasgow GI2 899, Scotland. U K Biol. 158, 567-572. 16. Parson, W.W. (1982) Photosynthetic bacterial reaction REFERENCES centres: interactions among the bacteriochlorophylls 1 Bachmann. R.C.. K. Gilliesand J.J. Takemoto (1981) and bacteriopheophytins. A n n u . Rev. Biophys. Bioeng. 11, 57-80. Membrane topography of the photosynthetic reaction 17. Peters, J. and G . Drews (1983) The lateral and c e n t r e p o l y p e p t i d e s of R h o d o p s e u d o m o n a s transverse topography of the membrane-bound pigsphaeroides. Biochemistry 20. 459&4596. 7 Brunisholz, R . A . , P.A. Cuendet. R. Theiler and H. ment-protein complexes of Rhodopseudonionas capsulara. In the Proceedings of the Workshop on Molecular Zuber (1981) The complete amino acid sequence of the Structure and Function of Light-Hanmtinp Pigmensingle light harvesting protein from chromatophores of Protein complexes and Photosynthetic Reaction CenRhodospirillum rubrum (3-9'. FEBS Lett. 129. 150tres, pp. 95. Zurich. 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(1983) The location. orientation and polypeptides (antenna complex B890) and of reaction stoichiometry of the Rieske iron-sulphur cluster in centre subunit L within the chromatophore membrane membranes of Rhodopseudomonas sphaeroides. from Rhodospirillum rubrum G-9'. Proceedings of the Biochim. Biophys. Acta 123, 133-138. Workshop on Molecular Srructure and Function of 21. Sutton, M . R . . D. Rosen. G. Feher and L . A . Steincr Lighi-harvesting Pigment-proiein Complexes and (1982) Amino-terminal sequences of the L. M and H Phoiosynthetic Reuctioti Centres. pp. 5 5 , Zurich. subunits of reaction centres from the photosynthetic Switzerland. bacterium Khodopseudomonas sphaeroides K-26. 5 Clayton, R.K. and W.R. Sistrom (1978) The PhoroBiochemistry 21. 3842-3849. rvnthetic Bacteria. Plenum Press. New York. 22. Tadros, M.H., F. Suter. G. Drewsand H . Zuber ( 1983) 6 Cogdell. R.J. and J . P . Thornber (1980) LightThe complete amino-acid sequence of the large bacterharvesting pigment-protein complexes of purple iochlorophyll-binding polypeptide from the lightphotosynthetic bacteria FEBS Leu. 122, 1-8. harvesting complex I1 (B80@850) of Rhodopseirdomo7 Cr0fts.A.R.. S . W . Meinhardt. and J.R. Bowyer(1982) nas capsulata. Eur. J . Biochem. 129, 533-536. The electron transport chain of Rhodopseudomonas 23. Theiler, R . and H. Zuber (1983) The light-harvesting .rphaeroides. In Functions of Quinones in Energy Conpolypeptides of Rhodopseudomonas sphueroides R serving Svstems. (Edited by B.L. Trumpower) pp. 26.l:II Conformational anal+ and the possible 447-498. Academic Press. New York. molecular structure of the hydrophobic domain o f the B 8 Cuendet. P.A., H . Zurrer. M. Snozzi and H. Zuber 850 complex. Eur. J . Biochem. ( I n press). (1978) On the localisation of a bacteriochlorophyll24. Theiler. R . , R. Brunisholz, G . Frank, F. 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tain cytochrome c,?Biochem. J . 189, 385-391. 33. Youvan, D.C.. M. Alberti. H . Begusch, E.J. Bylina and J.E. Hearst (1983) Reaction centre and lightharvesting I genes from Rhodopseudomonas capsulata. Proceedings of the Workshop on Molecular Structure and Function of Light-Hurvesfing Pigment-Protein complexes and Photosynthetic Reaction Centres. pp. 7 6 . Zurich. Switzerland. 34. Zinth. W . , W. Kaiser and H. Michel (1983) Efficient photochemical activity and strong dichroism of single crystals of reaction centres from Rhodopseudornonas viridis. Biochim. Eiophys. Actu. 723, 128-131. 35. Zuber. H.. R . A . Brunisholz, G. Frank. P. Fuglistaller. W. Sidler and R. Theiler (1983) Structure and arrangement of light-harvesting polypeptides and organisation of pigments in extramembrane (cyanobacteria) and intramembrane (photosynthetic bacteria) lightharvesting pigment-protein complexes. Is there a general structural principal for energy transfer? Considerations on the basis of the primary structure data of the polypeptides. In Proceedings of the Workshop on Molecular Structure and Function of Light-Harvesting Pigment-Protein complexes and Photosynthetic Reaction Centres. pp. 5 6 5 8 . Zurich, Switzerland.