Localization of the exposed N-terminal region of the B800-850 alpha and beta light-harvesting polypeptides on the cytoplasmic surface of Rhodopseudomonas capsulata chromatophores

Tadros, Monier H.; Frank, Rainer; Drews, Gerhart

doi:10.1128/jb.167.1.96-100.1986

Cited by 13 publications

(14 citation statements)

References 26 publications

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“…The results suggest that the N termini of the a polypeptides point to the cytoplasm but that they are protected from proteolytic attack on the cytoplasmic surface by proteinprotein or protein-phospholipid interaction. This idea is supported by the observation that in mutant strains of Rhodopseudomonas capsulata having only one of the LH complexes, B870 or B800-850, the a polypeptides are digested on the cytoplasmic surface of the membrane, but not in the wild-type strain of Rhodopseudomonas capsulata (27)(28)(29)(30). The P polypeptides only have short amino acid domains that extend the transmembrane a helices in the periplasmic side of the membrane.…”

Section: Discussionsupporting

confidence: 57%

“…The proteinase K-treated and untreated freeze-dried chromatophores and spheroplasts were extracted with organic solvent, and the extract was fractionated by column chromatography as described previously (27)(28)(29)(30).…”

Section: Resultsmentioning

confidence: 99%

“…Bacteriochlorophyll concentrations were calculated from absorption measurements of methanol-acetone (7:2, vol/vol) extracts at 770 nm with the absorption coefficient of 76 cm-l mM' (11) Pigment-binding polypeptides. The procedures for the isolation and purification of pigment-binding polypeptides from protease-treated and untreated membranes are described elsewhere (27)(28)(29)(30).…”

Section: Methodsmentioning

confidence: 99%

“…To understand the topography and organization of the pigment-protein complexes and the insertion and assembly of polypeptides in the membrane, the orientations of the N-and C-terminal domains and conserved residues in these regions are of importance. On the basis of protease degradation experiments and amino acid sequence analysis, a transmembrane orientation with the N termini facing the cytoplasm was postulated for B870a, B870p, and RC-L of Rhodospirillum rubrum (8,9) and for B800-850cx, B800-850p, B870a, B870p, and RC-L of Rhodopseudomonas capsulata (27)(28)(29) and R. sphaeroides (32). In this study, the orientations of the N and C termini of RC-L, RC-M, and LH polypeptides of R. sphaeroides were studied.…”

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Localization of reaction center and B800-850 antenna pigment proteins in membranes of Rhodobacter sphaeroides

et al. 1988

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The localization of the N-and C-terminal regions of pigment-binding polypeptides of the bacterial photosynthetic apparatus of Rhodobacter sphaeroides was investigated by proteinase K treatment of chromatophore and spheroplast-derived vesicles and amino acid sequence determination. Under conditions of proteinase K treatment of chromatophores, which left the in vivo absorption spectrum and the membrane intact, 15 and 46 amino acyl residues from the N-terminal regions of the L and M subunits, respectively, of the reaction center polypeptides were removed. The N termini are therefore exposed on the cytoplasmic surface of the membrane. The C-terminal domain of the light-harvesting B800-850a and B870a polypeptides was found to be exposed on the periplasmic surface of the membrane. A total of 9 and 13 amino acyl residues were cleaved from the B800-850a and B870a polypeptides, respectively, when spheroplasts were treated with proteinase K. The N-terminal regions of the a polypeptides were not digested in either membrane preparation and were apparently protected from proteolytic attack. Seven N-terminal amino acyl residues of the B800-850,1 polypeptide were removed after the digestion of chromatophores. C-terminal residues were not removed after the digestion of chromatophores or spheroplasts. The C termini seem to be protected from protease attack by interaction with the membrane. Therefore, the N-terminal regions of the , polypeptides are exposed on the cytoplasmic membrane surface. The C termini of the B polypeptides are believed to point to the periplasmic space.The photosynthetic apparatus of Rhodobacter sphaeroides is located in an intracytoplasmic membrane system formed by invaginations of the cytoplasmic membrane (15). The major components of the photosynthetic apparatus are the light-harvesting (LH) or antenna pigment-protein complexes and the photochemical reaction center (RC), which are present as integral membrane particles (14). The absorption of light energy by LH complexes creates excited singlet states which migrate over the antennae downhill to the RC where they are trapped and converted into a membrane potential difference and a redox potential difference. This conversion drives a cyclic electron transport and the formation of a proton gradient across the membrane (16).The bacteriochlorophyll and carotenoid pigments are bound in stoichiometric ratios to the polypeptides of the three complexes: RC and the LH complexes B870 and B800-850, named according to their near-infrared absorption bands. In each LH complex, two different polypeptides bind noncovalently two or three bacteriochlorophyll and one or two carotenoid molecules. The axes of the pigment molecules have a strict orientation relative to the plane of the membrane (6, 18, 25). The pigment-binding polypeptides span the membrane once by a transmembrane a helix of hydrophobic amino acyl residues in LH complexes (5). The RC subunits M and L (RC-M and RC-L, respectively) span the membrane five times each (1,13,22,39). The N-and C-terminal domains of L...

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Section: Discussionsupporting

confidence: 57%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Localization of reaction center and B800-850 antenna pigment proteins in membranes of Rhodobacter sphaeroides

et al. 1988

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“…An emerging theme is that the N termini of the complex polypeptides are exposed on or near the cytoplasmic surface of the photosynthetic membrane. This topographical relationship has been shown for the B800-850 and B870 a and 3 subunits and reaction center polypeptide L of Rhodopseudomonas capsulata (12,13), and B870 subunits and polypeptide L of Rhodospirillum rubrum (3), and now for the B875 a and, possibly, the a polypeptides of R. sphaeroides.…”

Section: Discussionmentioning

confidence: 97%

Transverse membrane topography of the B875 light-harvesting polypeptides of wild-type Rhodobacter sphaeroides

et al. 1987

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Purified B875 light-harvesting complex, chromatophores, and spheroplast-derived vesicles from wild-type Rhodobacter sphaeroides were treated with proteinase K or trypsin, and the a and ,B polypeptides were analyzed by electrophoretic, immunochemical, and protein-sequencing methods. With the purified complex, proteinase K digested both polypeptides and completely eliminated the A875 peak. Trypsin digested the a polypeptide and reduced the A875 by 50%. Proteinase K cleaved the , polypeptide of chromatophores and the a polypeptide of spheroplast-derived vesicles. Sequence analyses of polypeptides extracted from proteinase K-treated chromatophores revealed that the 18 polypeptide was cleaved between amino acids 4 and 5 from the N terminus. The N terminus of the a polypeptide was intact. We concluded that the N terminus of the 18 polypeptide is exposed on the cytoplasmic membrane surface, and the difference in the digestion patterns between the spheroplast-derived vesicles and chromatophores suggested that the C terminus of the a polypeptide is exposed on the periplasmic surface.The photosynthetic apparatus of Rhodobacter sphaeroides is found in an intracellular membrane system formed by invagination of the cytoplasmic membrane (6). Isolated membrane fractions contain several major polypeptides involved in the harvesting and use of light energy. The most abundant of these fractions are the bacteriochlorophyll aassociated polypeptides of the B800-850 and B875 lightharvesting complexes, which are named according to the wavelengths of light which they absorb (5, 6). These complexes harvest incident light and pass energy downhill to photochemical reaction centers. There, the energy is trapped and converted to an electrochemical potential across the membrane (6).The R. sphaeroides B875 complex contains two polypeptides, a and ,B, with 1:1 stoichiometry and molecular weights of 6,800 and 5,500, respectively (2, 18). Two moles each of bacteriochlorophyll a and carotenoid pigments are associated with these per each a and ,B polypeptide (2, 5). B875 complexes (and B870 complexes in related organisms) serve as energy transfer intermediaries between B800-850 complexes, carotenoids, and reaction centers (4,10,17). This role depends on the location and orientation of B875 in the lipid bilayer and its physicdl relationship to the other pigment-protein complexes.Comparatively little is known about the membrane topography of B875 in R. sphaeroides. Spectroscopic and membrane fusion studies indicate that B875 and B800-850 are laterally arranged in the membrane so that they are connected through the energy pathway (10, 17). Recently, the amino acid sequences of the a and a polypeptides have been determined, and this information has been used to speculate on the transverse arrangement of these polypeptides in the membrane (18). For example, in both polypeptides, a central domain containing a putative bacteriochlorophyll a binding site (histidine residue) and consisting of 20 hydrophobic amino acids is flanked by N-and C-terminal hydrop...

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Analysis of the Rhodobacter capsulatus puc operon: the pucC gene plays a central role in the regulation of LHII (B800-850 complex) expression.

et al. 1991

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Formation of the light harvesting complex B800‐850 (LHII) of Rhodobacter capsulatus requires the expression of more than the three known genes specific for that complex (pucA, pucB and pucE) encoding the alpha, beta and gamma subunits of LHII, respectively. In this work evidence is presented that the product of the gene pucC, which is located downstream from pucA, is essential for high‐level transcription of the pucBACDE operon and formation of LHII. Plasmids were constructed containing deletions in one or several puc genes and transferred to a pucC::Tn5 mutant in which the puc operon is not expressed. It was found that the LHII‐ phenotype of the mutant was due to the missing PucC protein and that all known puc genes are located in one operon. To dissect the pucC, pucD and pucE genes from pucB and pucA and independently regulate them, they were placed under control of the nifHDK promoter. Only under nitrogen‐fixing growth conditions was the LHII‐ pucC::Tn5 mutant complemented by this construction. It is concluded that expression of pucC is essential for formation of the LHII complex in R.capsulatus. Analysis of the pucD and pucE genes led to the conclusion that the products of these genes stabilize the B800‐850 complex.

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Localization of the exposed N-terminal region of the B800-850 alpha and beta light-harvesting polypeptides on the cytoplasmic surface of Rhodopseudomonas capsulata chromatophores

Cited by 13 publications

References 26 publications

Localization of reaction center and B800-850 antenna pigment proteins in membranes of Rhodobacter sphaeroides

Localization of reaction center and B800-850 antenna pigment proteins in membranes of Rhodobacter sphaeroides

Transverse membrane topography of the B875 light-harvesting polypeptides of wild-type Rhodobacter sphaeroides

Analysis of the Rhodobacter capsulatus puc operon: the pucC gene plays a central role in the regulation of LHII (B800-850 complex) expression.

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