Cross-Linking Studies on the Membrane Topography of Photosystem I Reaction Center Complex in Synechococcus sp.

The subunit arrangement of the photosystem I reaction centercomplex in the thylakoid membranes of the thermophilic cyanobacteriumSynechococcus sp. was examined using three cross-linking reagents.(1) Treatments of osmotically shocked and NaBr-washed protoplastswith low concentrations of hydrophilic cross-linking reagents,dimethyladipimidate and glutaraldehyde, preferentially decreased62, 60, 14 and 13 kDa polypeptides of the photosystem I reactioncenter complex resolved by SDS-polyacrylamide gel electrophoresis,together with the anchor protein and allophycocyanin which areassociated with the outer surface of the thylakoid membranes.This suggests that these four subunits of the photosystem Icomplex are exposed on the stromal surface of thylakoid membranes.In contrast, a hydrophobic cross-linker, hexamethylenediisocyanate,unspecifically cross-linked most of the membrane polypeptides.(2) The 13 and 14 kDa polypeptides decreased always in parallelto each other on treatment of the protoplasts or isolatd CP1-awith the three cross-linking reagents, and the disappearanceof the two polypeptides was accompanied by the appearance ofa cross-linked product(s), when fixed with glutaraldehyde andhexamethylenediisocyanate. The results suggest that the 13 and14 kDa polypeptides are neighboring polypeptides in the complex. (Received June 7, 1986; Accepted November 13, 1986)     

Assembly of Two Subunits of the Cyanobacterial Photosystem I on the n-Side of Thylakoid Membranes

Photosystem I contains several peripheral membrane proteins that are located on either positive (luminal) or negative (stromal or cytoplasmic) sides of thylakoid membranes of chloroplasts or cyanobacteria. Incorporation of two peripheral subunits into photosystem I of the cyanobacterium Synechocystis species PCC 6803 was studied using a reconstitution system in which radiolabeled subunits II (PsaD) and IV (PsaE) were synthesized in vitro and incubated with the isolated thylakoid membranes. After such incubation, the subunits were found in the membranes and were resistant to digestion with proteases and removal by 2 molar NaBr. All of the radioactive proteins incorporated in the membrane were found in the photosystem I complex. The subunit II was assembled specifically into cyanobacterial thylakoid membranes and not into Escherichia coli cell membranes or thylakoid membranes isolated from spinach. The assembly process did not require ATP or proton motive force, and it was not stimulated by ATP. The assembly of subunits II and IV into thylakoid membranes isolated from the strain AEK2, which lacks the gene psaE, was increased two- to threefold. The incorporation of subunit II was 15 to 17 times higher in the thylakoids obtained from the strain ADK3 in which the gene psaD has been inactivated. However, assembly of subunit IV in the same thylakoids was reduced by 65%, demonstrating that the presence of subunit II is required for the stable assembly of subunit IV. Large deletions in subunit II prevented its incorporation into thylakoids and assembly into photosystem I, suggesting that the overall conformation of the protein rather than a specific targeting sequence is required for its assembly into photosystem I.     

Topological Considerations of the 9-kDa Polypeptide which Contains Centers A and B, Associated with the 14- and 19-kDa Polypeptides in the Photosystem I Complex of Spinach

The topography of subunits around the 9-kDa polypeptide in thephotosystem I (PS I) complex of spinach was studied by examiningthe results of alkaline and chaotropic ion treatments, trypticdigestion, and cross-linking of thylakoid membranes supplementedwith Western blotting techniques using antibodies raised againstthe 9-, 14- and 19-kDa polypeptides. The 14- and 19-kDa polypeptideshave been shown to correspond to subunits III and II [Münchet al. (1988) Curr. Genet. 14: 511–518.], respectively,by a comparison of their respective amino acid compositionsand amino-terminal sequences [Oh-oka et al. (1988a) J. Biochem.103: 962–968.]. It appears that these three polypeptidesare peripheral proteins situated in close to each other on thestromal side of thylakoid membranes. The 9-kDa polypeptide withcenters A and B is stable within a specific environment of themembranes, in which the polypeptide is embedded under the twoother subunits, the 14- and 19-kDa polypeptides. Thus, the 9-kDapolypeptide becomes unstable when dissociated from the PS Icomplex and exposed to the solvent environment. (Received March 15, 1989; Accepted June 15, 1989)     

Organization of the thylakoid membrane from the heterotrophic cyanobacterium, Aphanocapsa 6714

The polypeptide composition of thylakoid membrane fractions from the heterotrophic cyanobacterium Aphanocapsa 6714 was examined by electrophoretic and immunoblotting procedures. We have identified thylakoid cytochromes f, b6, c-550 and c-553 by tetramethylbenzidine staining of lithium dodecyl sulfate polyacrylamide gels; we also have identified the Rieske Fe-S center protein and subunit 4 of the cytochrome b6/f complex. We have characterized phycobilisomes and active core preparations of PS I and PS II. PS I is comprised of five polypeptides (62 kDa, 14.5 kDa, 10 kDa, and two proteins of less than 10 kDa), and our PS II preparation is highly enriched for three chlorophyll-binding proteins of 48, 45 and 36 kDa. Furthermore, we have resolved the chlorophyll-binding complexes on non-denaturing gels and have determined the polypeptide composition of each chlorophyll-containing band. Three bands are associated with PS I (I, IIa and IIb) and three bands are PS II components (III', IIIa and IIIb) as judged by low-temperature fluorescence emission spectra. Band III' contains a 64 kDa antenna polypeptide, IIIa contains the 48 kDa and 45 kDa polypeptides, and IIIb is comprised solely of a 36 kDa protein. The IIIb apoprotein represents a novel PS II component; its possible role in photochemistry is discussed.     

Isolation and characterization of cDNA clones encoding photosystem I subunits with molecular masses 11.0, 10.0 and 8.4 kDa from Chlamydomonas reinhardtii

Summary cDNA clones encoding three photosystem I subunits of Chlamydomonas reinhardtii with apparent molecular masses 13, 5 and 3 kDa (thylakoid polypeptides 28, 35 and 37; P28, P35 and P37, respectively) were isolated using gene specific oligonucleotides as probes. The sequences of these oligonucleotides were deduced from the N-terminal amino acid sequences of the proteins. The cDNAs were sequenced and used to probe Southern and Northern blots. The Southern blot analysis indicates that the proteins are encoded by single-copy genes. The mRNA sizes of the three components are 960 (P28), 1120 (P35) and 790 (P37) nucleotides. Comparison between the open reading frames of the cDNAs and the N-terminal amino acid sequences of the proteins indicates that the nascent polypeptides possess N-terminal transit sequences that are removed to give mature proteins of 11.0 (P28), 10.0 (P35) and 8.4 (P37) kDa. Analysis of the deduced protein sequences suggests that P28 and P35 are extrinsic membrane proteins and that P37 spans the thylakoid membrane. All three proteins have short transit peptides that probably route them to the stromal side of the thylakoid membrane.Abbreviations OEE1, 2 and 3 oxygen evolution enhancer proteins 1, 2 and 3 - RuBisCO ribulose bisphosphate carboxylase/oxygenase - PS photosystem - P28, P35 and P37 Chlamydomonas reinhardtii thylakoid polypeptides 28, 35 and 37 The nucleotide sequences presented here will appear in the EMBL/Genbank/DDBJ Nucleotide Sequence Databases under the accession numbers X15164 (11.0 kDa subunit; P28), X15165 (10.0 kDa subunit; P35) and X15166 (8.4 kDa subunit; P37)     

Organization and Stability of Polypeptides Associated with the Chlorophyll a-b Light-Harvesting Complex of Photosystem-II

In the oxygen-evolving photosystem-II (PSII) of higher plantchioroplasts and green algae, most of the light-harvesting functionis performed by the chlorophyll (Chl) a-b-protein complex (LHC-II).On the average, the LHC-II contains about 210 Chl (a+b) moleculesper PSII reaction center. The polypeptide composition, copynumber and organization of assembly in the LHC-II complex arenot fully understood at present. This work utilized the chlorinaf2 mutant of barley (lacking Chl b and having a LHC-II antennaof only 13 Chl a molecules) to determine the organization andstability of assembly of proteins in the LHC-II. High-resolutionSDS-PAGE and immunoblot analysis showed the presence of fourmain constitutive polypeptides in the wild-type LHC-II (termedhere subunits a, b, c and d) with molecular masses in the range30–25 kDa. Of those, only subunit d (a 25 kDa polypeptide)was found to occur at an equal copy number per PSII reactioncenter in both wild-type and in the Chl b-less chlorina f2 mutant.All other subunits were either absent or existed in much loweramounts in the mutant. Subunit d is a polypeptide constituentof the major Chl-protein subcomplex (CPII) of the LHC-II. Itis stably incorporated in the thylakoid membrane in the absenceof Chl b and probably binds the 13 Chl a molecules in the residualLHC-II antenna of the chlorina f2 mutant. We propose that, ofall LHC-II polypeptides, subunit d is most proximal to the PSIIcore and may serve as a linker in the process of excitationenergy transfer from the bulk LHC-II to the PSII reaction centerin chloroplasts. (Received February 25, 1992; Accepted May 12, 1992)     

Photoactivation of Oxygen-Evolving Enzyme in Dark-Grown Pine Cotyledons: Relationship between Assembly of Photosystem II Proteins and Integration of Manganese and Calcium

Dark-grown cotyledons of pine (Pinus thunbergit) did not exhibitO₂ evolution, but this capability was rapidly activated by illuminationfor a short period (photoactivation). To examine the biochemicalchanges which accompany the process of photoactivation in gymnosperms,a method enabling the preparation of highly active O₂-evolvingphotosystem II (PS II) membranes was applied to light-grown,dark-grown, and photoactivated cotyledons. PS II membranes preparedfrom light-grown cotyledons exhibited high O₂-evolving activity,and contained all the intrinsic proteins as well as the threeextrinsic proteins (32, 23 and 17 kDa) associated with PS II.These membranes were also found to contain 4.4 Mn and 0.83 Ca/PSII reaction center. PS II membranes from dark-grown cotyledonscontained all the intrinsic proteins, but preserved only 32kDa extrinsic protein, and zero Mn and 0.85 Ca/PS II reactioncenter. The two extrinsic proteins (23 and 17 kDa) absent inthe PS II membranes from dark-grown cotyledons were, however,present as mature forms in whole thylakoid membranes from thecorresponding sample. The PS II membranes isolated from photoactivatedcotyledons showed a high activity of O₂ evolution and retainedthe three extrinsic proteins, 5.3 Mn and 1.1 Ca/PS II reactioncenter, respectively. The results indicated that Mn and thetwo extrinsic proteins were tightly integrated in the O₂-evolvingapparatusduring the process of photoactivation but integration of Capreceded the integration of Mn by photoactivation. (Received December 9, 1991; Accepted February 1, 1992)     

Glucolipid Synthesis Activities in Cytoplasmic and Thylakoid Membranes from the Cyanobacterium Anacystis nidulans

Cytoplasmic membranes (plasma membranes), thylakoid membranesand cell walls prepared from the cyanobacterium, Anacystis nidulans,were compared for UDP-glucose: l,2-diacylglycerol glucosyltransferaseactivity. When 1,2-dipalmitoylglycerol was added as a glucosylacceptor, both cytoplasmic membranes and thylakoid membranesincorporated glucose from UDP-glucose into monoglucosyl diacylglycerol,but the cell walls containing the outer membranes did not. Thecytoplasmic membranes incorporated about twice as much glucoseas the thylakoid membranes on a protein basis. These observationssuggest that in A. nidulans the UDP-glucose: 1,2-diacylglycerolglucosyltransferase participating in glucolipid biosynthesisis located in both cytoplasmic and thylakoid membranes, butnot in the outer membrane. ¹Solar Energy Research Group, The Institute of Physical andChemical Research (RIKEN), Wako-shi, Saitama 351-01, Japan. (Received November 21, 1985; Accepted January 27, 1986)     

Reversible and Irreversible Inactivation of Photosynthesis in Relation to the Lipid Phases of Membranes in the Blue-Green Algae (Cyanobacteria) Anacystis nidulans and Anabaena variabilis

The Arrhenius plots of photosynthetic oxygen evolution in theblue-green algae Anacystis nidulans and Anabaena variabiliswere composed of two straight lines with break points whichwere very close to temperatures for the onset of phase separationof the thylakoid membranes. Irreversible inactivation of photosynthesisbegan to appear at the same temperature as the onset of phaseseparation of the cytoplasmic membranes in A. nidulans. Electrolytesbegan to leak from the cytoplasm into the outer medium, indicatingthat the permeability of the cytoplasmic membranes increasedwhen they entered the phase separation state. In A. variabilis,in which the cytoplasmic membranes had remained in the liquidcrystalline state above 0?C, no irreversible damage to photosynthesisnor leakage of electrolytes was observed between 0 and 20?C.These findings suggest that photosynthesis of the blue-greenalgae is reversibly suppressed when only the thylakoid membranesare in the phase separation state, and irreversibly inactivatedwhen the cytoplasmic membranes are in the phase separation state. (Received April 9, 1984; Accepted June 19, 1984)     

Effect of heptane-extraction on the stability of subunit organization of photosystem I reaction center complexes

Treatment of lyophilized thylakoid membranes of the thermophilic cyanobacterium Synechococcus sp. with n-heptane for 6 h resulted in marked changes in the pattern of photosystem I reaction center complexes resolved by sodium dodecylsulfate-polyacrylamide gel electrophoresis. CP1-a, which consists of two large subunits and three small subunits, was a major chlorophyll-containing band resolved from the lyophilized thylakoid membranes, whereas the heptane-extracted membranes produced mainly CP1-e which totally lacks the small subunits. Electron transport from the primary donor P700 to the secondary acceptor P430 was not affected by the heptane-extraction of the membranes. The heptane-treatment removed 97% of -carotene present in the membranes, whereas all chlorophyll a, a major part of xanthophylls, more than a half of phylloquinone and one third of plastoquinone remained unextracted. The data suggest that -carotene has an important structural effect to stabilize the subunit organization of photosystem I reaction center complexes but is not essential for the early photochemical events of photosystem I.Abbreviations SDS sodium dodecylsulfate - PS photosystem     

Chlorophyll-Protein Complexes Associated with Photosystem I Isolated from the Green Alga, Bryopsis maxima

Six chlorophyll (Chl)-protein complexes associated with photosystemI (CPla), and the PS I reaction center complex (CPl) were isolatedfrom the thylakoid membranes of the green alga, Bryopsis maxima,by SDS-polyacrylamide gel electrophoresis. CPla had four polypeptides(22, 24, 25, 26 kDa) in addition to the 67 kDa polypeptide ofCPl. These complexes may thus possibly be a combination of CPland antenna complexes for PS I. Six CPla showed almost the sameoptical properties, with absorption maxima at 650 and 677 nmand contained carotene and a small amount of xanthophylls. TheChl a/b ratios of these CPla were about 2, while that of CPlwas 14. CPla showed a fluorescence emission maximum at 695 nm;its excitation spectrum had peaks at 438, 470 and 540 nm, correspondingto the absorption maxima of Chl a, Chl b, xanthophylls, respectively.An antenna complex free of CPl has been detected in some plantsbut was not found in the present alga. ¹Present address: Department of Botany, The University of Adelaide,Adelaide, S.A. 5001, Australia (Received April 17, 1986; Accepted June 26, 1986)     

Isolation and characterization of cDNA clones encoding the 17.9 and 8.1 kDa subunits of Photosystem I from Chlamydomonas reinhardtii

cDNA clones encoding two Photosystem I subunits of Chlamydomonas reinhardtii with apparent molecular masses of 18 and 11 kDa (thylakoid polypeptides 21 and 30; P21 and P30 respectively) were isolated using oligonucleotides, the sequences of which were deduced from the N-terminal amino acid sequences of the proteins. The cDNAs were sequenced and used to probe Southern and Northern blots. The Southern blot analysis indicates that both proteins are encoded by single-copy genes. The mRNA sizes of the two components are 1400 and 740 nucleotides, respectively. Comparison between the open reading frames of the cDNAs and the N-terminal amino acid sequences of the proteins indicates that the molecular masses of the mature proteins are 17.9 (P21) and 8.1 kDa (P30). Analysis of the deduced protein sequences predicts that both subunits are extrinsic membrane proteins with net positive charges. The amino acid sequences of the transit peptides suggest that P21 and P30 are routed towards the lumenal and stromal sides of the thylakoid membranes, respectively.Abbreviations OEE1, 2 and 3 oxygen evolution enhancer proteins 1, 2 and 3 - Rubisco ribulose bisphosphate carboxylase/oxygenase - PS photosystem - P21 and P30 C. reinhardtii thylakoid polypeptides 21 and 30     

Molecular Structure of a High-Potential Form of Thylakoid Cytochrome b-559 as Determined by Radiation Inactivation

Cytochrome b-559 in photosystem II can be characteristicallyconverted from a high- to a low-potential form. Taking thisresponse of Cyt b-559 as evidence for the denaturation of proteinmolecules, the sizes of the structures that stabilize the high-potentialform of Cyt b-559 in PS II membranes and thylakoids from spinachwere determined by radiation inactivation. When a target of26 kDa was inactivated in PS II membranes, Cyt b-559 was convertedto the low-potential form. The size was consistent with a molecularweight of Cyt b-559 in a proposed tetrameric structure thatconsists of two sets of 9.2-kDa and 4.3-kDa subunits [Widgeret al. (1985) FEBS Lett. 191: 186–190]. In contrast tothe functional size of 26 kDa in the PS II membranes, the functionalsize was 116 kDa in thylakoid membranes. The results suggestthe presence of an extra 90-kDa electron carrier between a redoxtitrator outside the membranes and the Cyt b-559, which maynot expose its active site to the surface of the thylakoids. (Received March 9, 1989; Accepted June 23, 1989)     

Protein rotational mobility in thylakoid membranes of different polypeptide composition in the wild type and mutant strains of Chlamydomonas reinhardtii

The rotational mobility of thylakoid membrane proteins labeled with a paramagnetic analog of N-ethylmaleimide was investigated by saturation transfer electron spin resonance. In the wild type strain of Chlamydomonas reinhardtii two polypeptides are prominently labeled. They correspond to the 19-kDa subunit of the reaction center I protein and to the 30-kDa subunit of the light harvesting complex. Several polypeptides, most of which are either trypsin or alkaline sensitive, are also labeled. In order to circumvent the lack of specificity during the labeling, we have compared the rotational mobilities of labeled proteins in thylakoid membranes from several mutant strains which lack in photosystem I., ATPase or light harvesting complexes. Comparison of the saturation transfer electron spin resonance spectra obtained with these mutant membranes as well as with trypsin- and alkaline-treated membranes allowed us to characterize the rotational contribution of some of the labeled proteins to the overall protein dynamics observed in the wild type strain. The reaction center I protein undergoes slow rotation as compared to the other labeled proteins. The rotational characteristics of the labeled light harvesting complexes are those of a peptide fragment in the complex which is in rapid motion in unstacked membranes. Stacking of the thylakoid membranes upon Mg²⁺ addition is accompanied by a marked change in shape of the saturation transfer spectra, and corresponds to the appearance of highly immobilized nitroxides. We interpret these changes as arising mainly from the hindrance upon membrane appression, of the labeled fragment of the light harvesting complexes which protrude at the thylakoid outer surface.     

Freezing damage and frost tolerance of the photosynthetic apparatus studied with isolated mesophyll protoplasts of Valerianella locusta L.

Mesophyll protoplasts were isolated from unhardened and cold-acclimated leaves of Valerianella locusta L. and subjected to freeze-thaw treatment. To evaluate the extent and course of freezing injury, photosynthetic reactions of whole protoplasts and of free thylakoid membranes, liberated from protoplasts by osmotic lysis, were measured. In addition, the integrity of the protoplasts was determined by microscopy. The results reveal an increased frost tolerance of protoplasts isolated from acclimated leaves with respect to all parameters measured. CO₂-dependent O₂ evolution (representing net photosynthetic CO₂ fixation of protoplasts) was the most freezing-sensitive reaction; its inhibition due to freeze-thaw treatment of protoplasts was neither correlated with disintegration of the plasma membrane, nor was it initiated by inactivation of the thylakoid membranes. The frost-induced decline of protoplast integrity was not closely correlated to thylakoid damage either. Freezing injury of the thylakoid membranes was manifested by inhibition of photosynthetic electron transport and photophosphorylation. Both photosystems were affected by freezing and thawing with strongest inhibition occurring in the water-oxidation system or at the oxidizing site of photosystem II. Photophosphorylation responded more sensitively to freezing stress than electron transport, although uncoupling (increased permeability of the thylakoid membranes to protons) was not a conspicuous effect. The data are discussed in relation to freezing injury in leaves and seem to indicate that frost damage in vivo is initiated at multiple sites.Abbreviations Chl chlorphyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DCIP 2,6-dichlorophenolindophenol - DPC 1,5-diphenylcarbazide - Hepes 2-[4-(2-hydroxyethyl)-1-piperazinyl]-ethanesulfonic acid - MES 2-(N-morpholino)-ethanesulfonic acid - PS I photosystem I - PS II photosystem II     

Extrinsic polypeptides of spinach photosystem I

By combining Triton X-114 partitioning with alkaline-salt and chaotropic washings of thylakoid membrane vesicles and photosystem I particles, we have studied the protein subunit composition and organization of spinach photosystem I. Upon fractionation of photosystem I particles with Triton X-114, 6 polypeptides of 5.0, 8.2 (psaE), 10.5, 16.6 (psaG), 19.3 and 22.1 kDa (psaD) were considered to be extrinsic membrane proteins. By combining this partitioning with salt washes of thylakoid membranes, the polypeptides of 8.2, 11.6 (psaH), 19.3 and 22.1 kDa were directly shown to be stromally oriented and extrinsic while no extrinsic subunits were identified at the inner thylakoid surface. The 5.0, 8.2, 10.5, 17.2, 19.3 and 22.1 kDa polypeptides appear to have regulatory rather than catalytic functions as their release from photosystem I particles upon high salt-alkali treatment does not affect photosystem I-mediated electron transport.Abbreviations DCIP 2.6-dichlorophenol indophenol - DCMU dichlorophenyl dimethyl urea - LHC light harvesting complex - PVDF polyvinylidene difluoride - SDS sodium dodecyl sulphate - TCA trichloroacetic acid - Tricine (N-tris[Hydroxymethyl]-methylglycine; N-[2-Hydroxy-1,1-bis(hydroxymethyl)-ethyl]glycine) - Tris (tris[Hydroxymethyl]aminomethane)     

Import of the barley PSI-F subunit into the thylakoid lumen of isolated chloroplasts

A full-length cDNA clone encoding the PSI-F subunit of barley photosystem I has been isolated and sequenced. The open reading frame encodes a precursor polypeptide with a deduced molecular mass of 24837 Da. The barley PSI-F precursor contains a bipartite presequence with characteristics similar to the presequences of proteins destined to the thylakoid lumen. In vitro import studies demonstrate that an in vitro synthesized precursor is transported across the chloroplast envelope and directed to the thylakoid membrane, where it accumulates in a protease-resistant form. Incubation of the precursor with a chloroplast stromal extract results in processing to a form intermediate in size between the precursor and mature forms. Hydrophobicity analysis of the barley PSI-F protein reveals a hydrophobic region predicted to be a membrane spanning -helix. The hydrophobic nature of PSI-F combined with a bipartite presequence is unusual. We postulate that the second domain in the bipartite presequence of the PSI-F precursor proteins is required to ensure the proper orientation of PSI-F in the thylakoid membrane. The expression of the PsaF gene is light-induced similar to other barley photosystem I genes.Abbreviations 16K 23K and 33K proteins, the 16 kDa, 23 kDa and 33 kDa subunits of the photosystem II oxygen-evolving complex - PSI-N and PSI-F photosystem I subunit N and F - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

Assembly and composition of the chlorophyll a-b light-harvesting complex of barley (Hordeum vulgare L.): Immunochemical analysis of chlorophyll b-less and chlorophyll b-deficient mutants

The chlorina-f2 mutant of barley (Hordeum vulgare L.) contains no chlorophyll b in its light-harvesting antenna, whereas the chlorina-103 mutant contains approximately 10% of the chlorophyll b found in wild-type. The absolute chlorophyll antenna size for Photosystem-II in wild-type, chlorina-103 and chlorina-f2 mutant was 250, 58 and 50 chlorophyll molecules, respectively. The absolute chlorophyll antenna size for Photosystem-I in wild-type, chlorina-103 and chlorina-f2 mutant was 210, 137 and 150 chlorophyll molecules, respoectively. In spite of the smaller PS I antenna size in the chlorina mutants, immunochemical analysis showed the presence of polypeptide components of the LHC-I auxiliary antenna with molecular masses of 25, 19.5 and 19 kDa. The chlorophyll a-b-binding LHC-II auxiliary antenna of PS II contained five polypeptide subunits in wild-type barley, termed a, b, c, d and e, with molecular masses of 30, 28, 27, 24 and 21 kDa, respectively. The polypeptide composition of the LHC-II auxiliary antenna of PS II was found to be identical in the two mutants, with only the 24 kDa subunit d present at an equal copy number per PS II in each of the mutants and in the wild-type barley. This d subunit assembles stably in the thylakoid membrane even in the absence of chlorophyll b and exhibits flexibility in its complement of bound chlorophylls. We suggest that polypeptide subunit d binds most of the chlorophyll associated with the residual PS II antenna in the chlorina mutants and that is proximal to the PS II-core complex.Abbreviations CP chlorophyll-protein - LHC the chlorophyll a-b binding light-harvesting complex - LHC-II subunit a the Lhcb4/5 gene product - subunit b the Lhcb1 gene product - subunit c Lhcb2 the gene product - subunit d the Lhcb3 gene product - subunit e the Lhcb6 gene product - PMSF phenylmethane sulphonyl fluoride - RC reaction center - Q_A the primary quinone electron acceptor of Photosystem-II - P700 the reaction center of PS I     

Outer membrane characteristics of Campylobacter jejuni grown in chickens

Abstract A type of in vivo phenotype of Campylobacter jejuni was obtained by maintaining bacteria in the peritoneal cavities of chickens for one week. These bacteria, which had not been subcultured on laboratory media, were used to prepare outer membranes for comparison with C. jejuni grown in vitro. Flagella with subunits of 65 kDa and a single porin with a protein subunit of 49 kDa were expressed constitutively; however, outer membrane proteins of 55, 35 and 20 kDa, and intermediate-chain lipopolysaccharide were only expressed by bacteria maintained in chickens.     

Is the 9 kDa thylakoid membrane phosphoprotein functionally and structurally analogous to the 'H' subunit of bacterial reaction centres?

Although the amino acid sequence of the 9 kDa (phospho)protein of chloroplasts has been determined, the function of this thylakoid membrane protein in photosynthetic electron transport and the reason for its physiological control remains unclear. In this paper, I briefly review the evidence which indicates that the phosphorylation of the 9 kDa protein results in a partial inhibition of photosynthetic oxygen evolution by increasing the stability of the semiquinone bound to QA the primary, plastoquinone-binding site of photosystem II (PS II). I propose that in its dephosphorylated state, the 9 kDa thylakoid membrane protein may serve PS II to ensure efficient photochemical charge separation by aiding the transfer of reducing equivalents out of the reaction centre to the attendant plastoquinone pool. This function is analogous to that proposed for the H-subunit of the reaction centre of photosynthetic eubacteria. Whether these two proteins have evolved from a common ancestral reaction centre protein is discussed in the light of a comparison of their amino acid sequences and predicted secondary structures.