植物光系统II放氧复合体外周蛋白结构和功能的研究进展

光合放氧是植物光系统II(PSII)的重要功能之一。PSII的放氧反应主要是由PSII氧化侧的 4个锰原子组成的锰簇催化的。在类囊体膜的囊腔侧还结合有若干个外周蛋白 ,对放氧反应起着重要作用。文章总结了植物光系统II外周蛋白的结构和功能研究方面的最新进展     

Mechanisms of phase behaviour and protein partitioning in detergent/polymer aqueous two-phase systems for purification of integral membrane proteins

Detergent/polymer aqueous two-phase systems are studied as a fast, mild and efficient general separation method for isolation of labile integral membrane proteins. Mechanisms for phase behaviour and protein partitioning of both membrane-bound and hydrophilic proteins have been examined in a large number of detergent/polymer aqueous two-phase systems. Non-ionic detergents such as the Triton series (polyoxyethylene alkyl phenols), alkyl polyoxyethylene ethers (C(m)EO(n)), Tween series (polyoxyethylene sorbitol esters) and alkylglucosides form aqueous two-phase systems in mixtures with hydrophilic polymers, such as PEG or dextran, at low and moderate temperatures. Phase diagrams for these mixtures are shown and phase behaviour is discussed from a thermodynamic model. Membrane proteins, such as bacteriorhodopsin and cholesterol oxidase, were partitioned strongly to the micelle phase, while hydrophilic proteins, BSA and lysozyme, were partitioned to the polymer phase. The partitioning of membrane protein is mainly determined by non-specific hydrophobic interactions between detergent and membrane protein. An increased partitioning of membrane proteins to the micelle phase was found with an increased detergent concentration difference between the phases, lower polymer molecular weight and increased micelle size. Partitioning of hydrophilic proteins is mainly related to excluded volume effects, i.e. increased phase component size made the hydrophilic proteins partition more to the opposite phase. Addition of ionic detergent to the system changed the partitioning of membrane proteins slightly, but had a strong effect on hydrophilic proteins, and can be used for enhanced separation between hydrophilic proteins and membrane protein.     

Structural organization of proteins on the oxidizing side of photosystem II. Two molecules of the 33-kDa manganese-stabilizing proteins per reaction center.

The 33-kDa manganese-stabilizing protein stabilizes the manganese cluster in the oxygen-evolving complex. There has been, however, a considerable amount of controversy concerning the stoichiometry of this photosystem II (PS II) component. In this paper, we have verified the extinction coefficient of the manganese-stabilizing protein by amino acid analysis, determined the manganese content of oxygen-evolving photosystem II membranes and reaction center complex using inductively coupled plasma spectrometry, and determined immunologically the amount of the manganese-stabilizing protein associated with photosystem II. Oxygen-evolving photosystem II membranes and reaction center complex preparations contained 258 +/- 11 and 67 +/- 3 chlorophyll, respectively, per tetranuclear manganese cluster. Immunoquantification of the manganese-stabilizing protein using mouse polyclonal antibodies on "Western blots" demonstrated the presence of 2.1 +/- 0.2 and 2.0 +/- 0.3 molecules of the manganese-stabilizing protein/tetranuclear manganese cluster in oxygen-evolving PS II membranes and highly purified PS II reaction center complex, respectively. Since the manganese-stabilizing protein co-migrated with the D2 protein in our electrophoretic system, accurate immunoquantification required the inclusion of CaCl2-washed PS II membrane proteins or reaction center complex proteins in the manganese-stabilizing protein standards to compensate for the possible masking effect of the D2 protein on the binding of the manganese-stabilizing protein to Immobilon-P membranes. Failure to include these additional protein components in the manganese-stabilizing protein standards leads to a marked underestimation of the amount of the manganese-stabilizing protein associated with these photosystem II preparations.     

Structural analysis of the hepatic glucagon receptor. Identification of a guanine nucleotide-sensitive hormone-binding region

[125I-Tyr10]Monoiodoglucagon [( 125I]MIG) was cross-linked to liver membrane glucagon receptors with hydroxysuccinimidyl-p-azidobenzoate, and the products were analyzed by sodium dodecyl sulfate-gel electrophoresis. Autoradiograms of the gel obtained after a 24-h exposure showed one major band at Mr = 63,000 that was sensitive to GTP and excess unlabeled glucagon. Exposure for 7 days showed labeling of an additional Mr = 33,000 species that was also sensitive to excess unlabeled glucagon. The Mr = 33,000 peptide can be obtained by subtilisin, trypsin, elastase, or Staphylococcus aureus V8 protease treatment of the [125I]MIG-occupied receptor in the membrane or in Lubrol-PX solution. In contrast, limited proteolysis of membranes containing vacant receptors results in labeling of a Mr = 24,000 peptide. The Mr = 24,000 peptide specifically binds [125I]MIG in a GTP-sensitive manner. The Mr = 33,000 peptide also retains GTP sensitivity since it releases bound [125I]MIG upon addition of GTP. Elastase treatment of the electroeluted Mr = 33,000 peptide yields the Mr = 24,000 and 15,000 fragments. The Mr = 15,000 peptide is the smallest fragment of the receptor as yet identified. Treatment of the Mr = 63,000 receptor with [125I]MIG cross-linked to it with endo-beta-N-acetylglucosaminidase F results in four distinct fragments with Mr values of 61,000, 56,000, 51,000, and 45,000; prolonged treatment resulted in the accumulation of the last two. Neither the Mr = 33,000 nor the Mr = 24,000 fragment appeared to be substrates for endo-beta-N-acetylglucosaminidase F. These data indicate that glucagon receptor is a glycoprotein of approximately 60,000 daltons which contains at least four N-linked glycans accounting for 18,000 daltons of its mass. Both its glucagon binding function and its capacity to interact with the stimulatory regulator of adenylyl cyclase are contained within a fragment of only approximately 21,000 daltons that does not contain any N-linked glycans. Hormone occupancy of the receptor results in a conformational change so as to expose a region that is susceptible to proteolysis by proteases of varying specificities to yield a peptide of approximately 30,000 daltons that also does not contain N-linked glycans.     

Leaf senescence in a non-yellowing mutant of Festuca pratensis: Proteins of photosystem II

The senescence of leaves is characterized by yellowing as chlorophyll pigments are degraded. Proteins of the chloroplasts also decline during this phase of development. There exists a non-yellowing mutant genotype of Festuca pratensis Huds. which does not suffer a loss of chlorophyll during senescence. The fate of chloroplast membrane proteins was studied in mutant and wild-type plants by immune blotting and immuno-electron microscopy. Intrinsic proteins of photosystem II, exemplified by the light-harvesting chlorophyll a/b-binding protein (LHCP-2) and D1, were shown to be unusually stable in the mutant during senescence, whereas the extrinsic 33-kilodalton protein of the oxygen-evolving complex was equally lable in both genotypes. An ultrastructural study revealed that while the intrinsic proteins remained in the internal membranes of the chloroplasts, they ceased to display the heterogenous lateral distribution within the lamellae which was characteristic of nonsenescent chloroplasts. These observations are discussed in the light of possible mechanisms of protein turnover in chloroplasts.Abbreviations kDa kilodalton - LHCP-2 light-harvesting chlorophyll a/b-binding protein - M_r relative molecular mass - PSII photosystem II - SDS sodium dodecyl sulphate     

Polypeptides of photosystem II and their role in oxygen evolution

The linear, four-step oxidation of water to molecular oxygen by photosystem II requires cooperation between redox reactions driven by light and a set of redox reactions involving the S-states within the oxygen-evolving complex. The oxygenevolving complex is a highly ordered structure in which a number of polypeptides interact with one another to provide the appropriate environment for productive binding of cofactors such as manganese, chloride and calcium, as well as for productive electron transfer within the photoact. A number of recent advances in the knowledge of the polypeptide structure of photosystem II has revealed a correlation between primary photochemical events and a core complex of five hydrophobic polypeptides which provide binding sites for chlorophyll a, pheophytin a, the reaction center chlorophyll (P680), and its immediate donor, denoted Z. Although the core complex of photosystem II is photochemically active, it does not possess the capacity to evolve oxygen. A second set of polypeptides, which are water-soluble, have been discovered to be associated with photosystem II; these polypeptides are now proposed to be the structural elements of a special domain which promotes the activities of the loosely-bound cofactors (manganese, chloride, calcium) that participate in oxygen evolution activity. Two of these proteins (whose molecular weights are 23 and 17 kDa) can be released from photosystem II without concurrent loss of functional manganese; studies on these proteins and on the membranes from which they have been removed indicate that the 23 and 17 kDa species from part of the structure which promotes retention of chloride and calcium within the oxygen-evolving complex. A third water-soluble polypeptide of molecular weight 33 kDa is held to the photosystem II core complex by a series of forces which in some circumstances may include ligation to manganese. The 33 kDa protein has been studied in some detail and appears to promote the formation of the environment which is required for optimal participation by manganese in the oxygen evolving reaction. This minireview describes the polypeptides of photosystem II, places an emphasis on the current state of knowledge concerning these species, and discusses current areas of uncertainty concerning these important polypeptides.Abbreviations A 23187 ionophore that exchanges divalent cations with H⁺ - Chl chlorophyll - cyt cytochrome - DCPIP dichlorophenolindophenol - DPC diphenylcarbazide - EGTA ethyleneglycoltetraacetic acid - P680 the chlorophyll a reaction center of photosystem II - pheo pheophytin - PQ plastoquinone - PS photosystem - Q_A and Q_B primary and secondary plastoquinone electron acceptors of photosystem II - Sn (n=0, 1, 2, 3, 4) charge accumulating state of the oxygen evolving system - Signals II_vf, II_f and II_s epr-detectable free radicals associated with the oxidizing side of photosystem II - Z primary electron donor to the photosystem II reaction center The survey of literature for this review ended in September, 1984.     

The topology of a membrane protein: the orientation of the 32 kd Qb-binding chloroplast thylakoid membrane protein

Exposed portions of the 32 kd chloroplast membrane quinone-binding and triazine herbicide-binding protein of photosystem II have been mapped to the lumenal or to the outer (stromal) surface of the thylakoid by following reactions of antibodies generated against synthetic peptides corresponding to predicted hydrophilic amino acid sequences with normally oriented or everted membrane vesicles. These data have led to the construction of a model with five membrane-spanning domains. The model has been verified, in part, by immunoblots of fragments of the protein produced by trypsin treatment of thylakoids with peptide-specific antibodies. Some of the hydrophilic loops appear to be in close contact with proteins of the oxygen evolving complex of photosystem II inasmuch as their removal increases the antibody reaction.     

A light-dependent mechanism for massive accumulation of manganese in the photosynthetic bacterium Synechocystis sp. PCC 6803

Manganese is an essential micronutrient for many organisms. Because of its unique role in the water oxidizing activity of photosystem II, manganese is required for photosynthetic growth in plants and cyanobacteria. Here we report on the mechanism of manganese uptake in the cyanobacterium Synechocystis sp. PCC 6803. Cells grown in 9 microM manganese-containing medium accumulate up to 1 x 10(8) manganese atoms/cell, bound to the outer membrane (pool A). This pool could be released by EDTA treatment. Accumulation of manganese in pool A was energized by photosynthetic electron flow. Moreover, collapsing the membrane potential resulted in the immediate release of this manganese pool. The manganese in this pool is mainly Mn(II) in a six-coordinate distorted environment. A distinctly different pool of manganese, pool B ( approximately 1.5 x 10(6) atoms/cell), could not be extracted by EDTA. Transport into pool B was light-independent and could be detected only under limiting manganese concentrations (1 nM). Evidently, manganese uptake in Synechocystis 6803 cells occurs in two steps. First, manganese accumulates in the outer membrane (pool A) in a membrane potential-dependent process. Next, manganese is transported through the inner membrane into pool B. We propose that pool A serves as a store that allows the cells to overcome transient limitations in manganese in the environment.     

Composition of major outer membrane proteins of Vibrio vulnificus isolates: effect of different growth media and iron deficiency

The outer membrane proteins of Vibrio vulnificus including isolates from humans, seawater and an asari clam were examined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A major outer membrane protein with an apparent molecular weight of 48,000 (48K protein) was common to all the strains grown in 3% NaCl-nutrient broth; however this 48K protein was not produced in any of the strains grown in chemically defined medium. Other major outer membrane proteins with molecular weights ranging from 33,000 to 40,000 varied in number, relative amount and molecular weight depending on the strain. One to three new outer membrane proteins with molecular weights ranging from 74,000 to 85,000 were produced in the cells grown in iron-deficient medium. The 48K protein and one or two major proteins with molecular weights ranging from 35,000 to 37,000 in the cells grown in 3% NaCl-nutrient broth were not solubilized by 2% SDS at 60 C for 30 min and were resistant to trypsin, indicating that they are porins. On the other hand, in cells grown in chemically defined medium, one or two major outer membrane proteins with molecular weights ranging from 33,000 to 40,000 might be porins.     

Quantitative analysis of membrane components in a highly active O2-evolving photosystem II preparation from spinach chloroplasts

Stoichiometry of membrane components associated with Photosystem II was determined in a highly active O₂-evolving Photosystem II preparation isolated from spinach chloroplasts by the treatment with digitonin and Triton X-100. From the analysis with sodium dodecyl sulfate polyacrylamide gel electrophoresis and Triton X-114 phase partitioning, the preparation was shown to contain the reaction center protein (43 kDa), the light-harvesting chlorophyll-protein complex (the main band, 27 kDa), the herbicide-binding protein (32 kDa) and cytochrome b-559 (10 kDa) as hydrophobic proteins, and three proteins (33, 24 and 18 kDa) which probably constitute the O₂-evolution enzyme complex as hydrophilic proteins. These proteins were associated stoichiometrically with the Photosystem II reaction center: one Photosystem II reaction center, approx. 200 chlorophyll, one high-potential form of cytochrome b-559, one low-potential form of cytochrome b-559, one 33 kDa protein, one (to two) 24 kDa protein and one (to two) 18 kDa protein. Measurement of fluorescence induction showed the presence of three electron equivalents in the electron acceptor pool on the reducing side of Photosystem II in our preparation. Three molecules of plastoquinone A were detected per 200 chlorophyll molecules with high-performance liquid chromatography. The Photosystem II preparation contained four managanese atoms per 200 chlorophyll molecules.     

Highly efficient purification of the 33-, 24-, and 18-kDa proteins in spinach photosystem II by butanol/water phase partitioning and high-performance liquid chromatography

The 33-, 24-, and 18-kDa proteins involved in photosynthetic oxygen evolution were purified from spinach photosystem II particles by butanol/water phase partitioning and high-performance liquid chromatography with a silica-based cation-exchange column. With this procedure a significant improvement was made in the time required for the purification and also in the amount and purity of the proteins. The N-terminal sequence of amino acid was determined for the purified proteins. Partial degradation of the proteins, which sometimes occurred in the purification, was not detected in the new procedure.     

Characterization of the Hepatic Glucagon Receptor

Abstract

The hepatic glucagon receptor was covalently labeled with [¹²⁵I-Tyr¹⁰]-monoiodoglucagon by use of the heterobifunctional crosslinker hydroxysuccini-midyl-p-azidobenzoate and analyzed by SDS-gel electrophoresis. The autoradio-gram of the gel showed one band at M_r=63,000 that was sensitive to excess unlabeled glucagon and GTP. The labeled receptor was solubilized with Lubrol-PX and the hydrodynamic characteristics of the receptor were determined. The molecular parameters of the solubilized receptor are S_{20, w} = 4.3 ± 0.1, Stokes radius = 6.3 ± 0.1 nm, frictional coefficient f/f° = 1.8 and a calculated M_r = 33,000 fragment, that retains guanine nucleotide sensitivity. Elastase treatment of vacant receptors results in a M_r = 24,000 fragment that binds hormone in a GTP-sensitive manner. The M_r = 24,000 fragment is contained within the M_r = 33,000 fragment. The M_r = 63,000 receptor upon treatment with endo-β-N-acetylglucosamine F for 4 h yields four fragments of apparent M_r = 61,000, 56,000, 51,000, and 45,000; 24 h treatment results in the accumulation of the last two fragments. Neither M_r = 33,000 and 24,000 fragment appear to be substrates for endo-β-N-acetylglucosaminidase F.

These data allow us to conclude that the hepatic glucagon receptor in the membrane is a dimer of ～ 60,000 dalton hormone binding subunit which is a glycoprotein containing at least four N-linked glycans accounting for 18,000 daltons of its mass. Both the hormone binding function and the capacity for the interaction with the stimulatory regulator of adenylyl cyclase are contained within a fragment of only ～ 21,000 daltons that does not contain any N-linked glycans.     

Evidence for thylakoid membrane fusion during zygote formation in Chlamydomonas reinhardtii

To understand whether fusions of thylakoid membranes from the parental chloroplasts occurred during zygote formation in Chlamydomonas reinhardtii, we performed an ultrastructural analysis of the zygotes produced by crossing mutants lacking photosystem I or II protein complexes, in the absence of de novo chloroplast protein synthesis. Thylakoid membranes from each parent could be distinguished on thin sections due to their organization in "supergrana" in mutants lacking photosystem I centers, by freeze-fracturing due to the absence of most of the exoplasmic-face (EF) particles in mutants lacking photosystem II centers, by immunocytochemistry using antibodies directed against photosystem II subunits. We demonstrate that a fusion of the thylakoid membranes occurred during zygote formation approximately 15 h after mating. These fusions allowed a lateral redistribution of the thylakoid membrane proteins. These observations provide the structural basis for the restoration of photosynthetic electron flow in the mature zygote that we observed in fluorescence induction experiments.     

Identification of three previously unknown in vivo protein phosphorylation sites in thylakoid membranes of Arabidopsis thaliana

The proteins in plant photosynthetic thylakoid membranes undergo light-induced phosphorylation, but only a few phosphoproteins have been characterized. To access the unknown sites of in vivo protein phosphorylation the thylakoid membranes were isolated from Arabidopsis thaliana grown in normal light, and the surface-exposed peptides were cleaved from the membranes by trypsin. The peptides were methylated and subjected to immobilized metal affinity chromatography, and the enriched phosphopeptides were sequenced using tandem nanospray quadrupole time-of-flight mass spectrometry. Three new phosphopeptides were revealed in addition to the five known phosphorylation sites in photosystem II proteins. All phosphopeptides are found phosphorylated at threonine residues implementing a strict threonine specificity of the thylakoid kinases. For the first time protein phosphorylation is found in photosystem I. The phosphorylation site is localized to the first threonine in the N terminus of PsaD protein that assists in the electron transfer from photosystem I to ferredoxin. A new phosphorylation site is also revealed in the acetylated N terminus of the minor chlorophyll a-binding protein CP29. The third novel phosphopeptide, composed of 25 amino acids, belongs to a nuclear encoded protein annotated as "expressed protein" in the Arabidopsis database. The protein precursor has a chloroplast-targeting peptide followed by the mature protein with two transmembrane helices and a molecular mass of 14 kDa. This previously uncharacterized protein is named thylakoid membrane phosphoprotein of 14 kDa (TMP14). The finding of the novel phosphoproteins extends involvement of the redox-regulated protein phosphorylation in photosynthetic membranes beyond the photosystem II and its light-harvesting antennae.     

Comparative studies on actins from various sources. Fragments of actins from Ascaris muscle cleaved at cysteinyl residues in comparison with those of other actins.

Pure actins were obtained from various animal muscles: Vertebrata (skeletal, smooth, and cardiac muscles), Prochordata (smooth muscle), Nematoda (obliquely striated muscle), and Mollusca (striated, smooth and obliquely striated muscles). These actins were all identical in apparent molecular weight on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. All actins treated with 2-nitro-5-thiocyanobenzoic acid yielded four major (about 33,000, 26,000, 24,000, and less than 10,000 daltons) and three minor (22,000, 17,000, and 10,000 daltons) bands in addition to intact actin on gel electrophoresis. The results suggest that all actins from various types of muscle have cysteinyl residues at similar positions on the primary structure.     

Calcium retards NH2OH inhibition of O2 evolution activity by stabilization of Mn2+ binding to photosystem II

Calcium is required for oxidation of water to molecular oxygen by photosystem II; the Ca2+ demand of the reaction increases upon removal of 23- and 17-kDa extrinsic polypeptides from detergent-derived preparations of the photosystem. Employing the manganese reductant NH2OH as a probe to examine the function of Ca2+ in photosystem II reveals that (1) Ca2+ slows the rate of NH2OH inhibition of O2 evolution activity, but only in photosystem II membranes depleted of extrinsic proteins, (2) other divalent cations (Sr2+, Cd2+) that compete for the Ca2+ site also slow NH2OH inhibition, (3) Ca2+ is noncompetitive with respect to NH2OH, (4) in order to slow inhibition, Ca2+ must be present prior to the initiation of NH2OH reduction of manganese, and (5) Ca2+ appears not to interfere with NH2OH reduction of manganese. We conclude that the ability of Ca2+ to slow the rate of NH2OH inhibition arises from the site in photosystem II where Ca2+ normally stimulates O2 evolution and that the mechanism of this phenomenon arises from the ability of Ca2+ or certain surrogate metals to stabilize the ligation environment of the manganese complex.     

Localization of 13 one-helix integral membrane proteins in photosystem II subcomplexes

Photosystem II is a multimeric protein complex of the thylakoid membrane in chloroplasts. Approximately half of the at least 26 different integral membrane protein subunits have molecular masses lower than 10 kDa. After one-dimensional (1D) or two-dimensional (2D) polyacrylamide gel electrophoresis (PAGE) separation, followed by enzymatic digestion of detected proteins, hardly any of these low-molecular-weight (LMW) subunits are detectable. Therefore, we developed a method for the analysis of highly hydrophobic LMW proteins. Intact proteins are extracted from acrylamide gels using a mixture of formic acid and organic solvent, precipitated with acetone, and analyzed by “top-down” mass spectrometry (MS). After offline nanoESI (electrospray ionization) MS, all LMW one-helix proteins from photosystem II were detected. In the four detected photosystem II supercomplexes of Nicotiana tabacum wild-type plants, 11 different one-helix proteins were identified as PsbE, -F, -H, -I, -K, -L, -M, -Tc, -W, and two isoforms of PsbX. The proteins PsbJ, -Y1, and -Y2 were localized in the buffer front after blue native (BN) PAGE, indicating their release during solubilization. Assembled PsbW is detected exclusively in supercomplexes, whereas it is absent in photosystem II core complexes, corroborating the protein’s function for assembly of the light-harvesting complexes. This approach will substantiate gel-blot immunoanalysis for localization and identification of LMW protein subunits in any membrane protein complex.     

Biogenesis of chloroplast membranes: XIV. Inhomogeneity of membrane protein distribution in photosystem particles obtained from Chlamydomonas reinhardi. Y-l

Treatment of chloroplast membranes of Chlamydomonas reinhardi with Triton-× 100 yielded membrane particles which were resolved into three bands on discontinuous sucrose gradients. One of these was enriched in the chlorophyll absorption and fluorescence properties and photosynthetic activities consistent with photosystem I enrichment, while another had the chlorophyll absorption and fluorescence properties expected to photosystem II enriched particles. The third type of particle was enriched in chlorophyll species which are probably the bulk chlorophylls of photosystem I. Analysis of the proteins of these fractions by polyacrylamide electrophoresis indicated substantial differences, the most striking being that the photosystem II particle type was greatly enriched in the major species of chloroplast membrane protein. Previous work has shown this to be an important protein controlling membrane assembly. This protein was depleted in the photosystem I particle type. We interpret this data to indicate a lack of homogeneity in the distribution of membrane proteins in the chloroplast membranes of Chlamydomonas, at the level of the two photosystems.     

Photosynthetic activity and membrane polypeptide composition of supergranal chloroplasts from plant tissue cultures containing a viruslike particle

Tissue culture cells of Streptanthus tortuosus (Kell.) var. orbiculatus (Greene) Hall (Cruciferae), having a viruslike particle in their nucleoli, the STV cell line, contain “supergranal” chloroplasts. Freeze-fracture studies of chloroplasts of a control cell line, which lacks the viruslike particles, reveal two complementary faces similar to those observed in spinach chloroplasts. Replicas of freeze-fractured STV supergranal chloroplasts, however, show that one membrane face (B) contains widely spaced 80 Å particles and the other face (C) is essentially smooth. Isolated STV supergranal chloroplasts lack photosystem II activity as indicated by their inability to reduce dichlorophenolindophenol and are unable to reduce NADP with electrons from photosystem II or from ascorbate-reduced dichlorophenolindophenol. However, partial photosystem I activity is indicated by the reduction of methyl viologen with electrons from dichlorophenolindophenol-ascorbate. This supports the concept that there is not a direct correspondence between grana formation and photosystem II activity. Electrophoresis shows that all of the major polypeptide bands present in the STV supergranal chloroplasts are also present in the control chloroplast membranes. One band, molecular weight 33,000, is present in a greatly increased amount in the STV supergranal chloroplast membranes and may be associated with grana stacking.