Isolation and Characterization of the Central Component of the Phycobilisome Core of Nostoc sp

Freshly isolated allophycocyanin is recovered from linear sucrose gradients made in 0.75 molar potassium phosphate buffer (pH 7.0) in three sizes: 19s, 10.3s, and 5.5s. The largest aggregate is a complex of a 680 nm fluorescing allophycocyanin I in the form (αβ)₃γ, where γ is the 95 kilodalton (kD) polypeptide, and two 660 nanometer fluorescing allophycocyanin II (αβ)₃ molecules; the complex, stabilized in high phosphate concentrations, fluoresces maximally at 675 nanometers. The 10.3s fraction is a hexamer of allophycocyanin of the 660 nanometer fluorescing type, perhaps attached through two polypeptides of 46 kD and 44 kD. The 5.5s component of the allophycocyanin pool is the usual trimeric form of allophycocyanin (αβ)₃. A similar 19s fraction is the major component of allophycocyanin I isolated under optimum conditions in the presence of the protease inhibitor, phenylmethylsulfonylfluoride. This 19s fraction is apparently a central component of the core of the phycobilisome with its 95 kD polypeptide the attachment point of the phycobilisome and membrane. The 95 kD polypeptide has both long wavelength absorption and fluorescence bands which seem to account for the long wavelength fluorescence properties of allophycocyanin I.     

Constant Phycobilisome Size in Chromatically Adapted Cells of the Cyanobacterium Tolypothrix tenuis, and Variation in Nostoc sp

Phycobilisomes of Tolypothrix tenuis, a cyanobacterium capable of complete chromatic adaptation, were studied from cells grown in red and green light, and in darkness. The phycobilisome size remained constant irrespective of the light quality. The hemidiscoidal phycobilisomes had an average diameter of about 52 nanometers and height of about 33 nanometers, by negative staining. The thickness was equivalent to a phycocyanin molecule (about 10 nanometers). The molar ratio of allophycocyanin, relative to other phycobiliproteins always remained at about 1:3. Phycobilisomes from red light grown cells and cells grown heterotrophically in darkness were indistinguishable in their pigment composition, polypeptide pattern, and size. Eight polypeptides were resolved in the phycobilin region (17.5 to 23.5 kilodaltons) by isoelectric focusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Half of these were invariable, while others were variable in green and red light. It is inferred that phycoerythrin synthesis in green light resulted in a one for one substitution of phycocyanin, thus retaining a constant phycobilisome size. Tolypothrix appears to be one of the best examples of phycobiliprotein regulation with wavelength. By contrast, in Nostoc sp., the decrease in phycoerythrin in red light cells was accompanied by a decrease in phycobilisome size but not a regulated substitution.     

Differentiation between Phycobiliprotein and Colorless Linker Polypeptides by Fluorescence in the Presence of ZnSO(4)

Microcystis aeruginosa, a unicellular cyanobacterium, contains small phycobilisomes consisting of C-phycocyanin, allophycocyanin, and linker polypeptides. SDS-polyacrylamide gels of the phycobilisomes were examined for fluorescent bands before and after spraying with a solution of ZnSO₄, followed by Coomassie brilliant blue staining for protein. This procedure provides a rapid and sensitive method for detecting small amounts of phycobilin-containing polypeptides and distinguishing them from other tetrapyrrole-containing polypeptides and from `colorless' ones. Three polypeptide bands, in addition to the α and β phycobiliprotein subunits, have been detected under these conditions. An 85 kilodalton polypeptide was identified as a phycobiliprotein due to its enhanced fluorescence in the presence of ZnSO₄. The other polypeptides do not contain chromophores and are colorless. They are approximately 34.5 and 30 kilodaltons in size.     

Chlorophyll-Protein Complexes of the Cyanophyte, Nostoc sp

Four chlorophyll-protein complexes have been resolved from the cyanophyte, Nostoc sp., by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis at 4 C. Complexes solubilized by SDS from Spinacia oleracea were run for comparison. As has been well documented, the P700-chlorophyll a-protein complex from the higher plant and blue-green algal samples are similar, and the light-harvesting pigment protein complex is present only in the former. Most noteworthy are two closely migrating chlorophyll proteins in Nostoc sp. which have approximately the same mobility as a single chlorophyll-protein band resolvable from spinach. The absorption maximum of the complex from spinach is at 667 nanometers, and those of the two complexes from Nostoc sp. are at 667 and 669 nanometers; the fluorescence emission maximum at −196 C is at 685 nanometers, and the 735 nanometer fluorescence peak, characteristic of the P700-chlorophyll a-protein complex, is absent. The apoproteins of these new complexes from Nostoc sp. and spinach are in the kilodalton range. It appears that at least one of these two chlorophyll-protein complexes from Nostoc sp. compares with those recently described by others from higher plants and green algae as likely photosystem II complexes, perhaps containing P680, although no photochemical data are yet available.     

Phycobilisome linker proteins are phosphorylated in Synechocystis sp. PCC 6803

The controversial issue of protein phosphorylation from the photosynthetic apparatus of Synechocystis sp. PCC 6803 has been reinvestigated using new detection tools that include various immunological and in vivo labeling approaches. The set of phosphoproteins detected with these methods includes ferredoxin-NADPH reductase and the linker proteins of the phycobilisome antenna. Using mutants that lack a specific set of linker proteins and are affected in phycobilisome assembly, we show that the phosphoproteins from the phycobilisomes correspond to the membrane, rod, and rod-core linkers. These proteins are in a phosphorylated state within the assembled phycobilisomes. Their dephosphorylation requires partial disassembly of the phycobilisomes and further contributes to their complete disassembly in vitro. In vivo we observed linker dephosphorylation upon long-term exposure to higher light intensities and under nitrogen limitation, two conditions that lead to remodeling and turnover of phycobilisomes. We conclude that this phosphorylation process is instrumental in the regulation of assembly/disassembly of phycobilisomes and should participate in signaling for their proteolytic cleavage and degradation.     

Phycobilisome Mobility and Its Role in the Regulation of Light Harvesting in Red Algae

Red algae represent an evolutionarily important group that gave rise to the whole red clade of photosynthetic organisms. They contain a unique combination of light-harvesting systems represented by a membrane-bound antenna and by phycobilisomes situated on thylakoid membrane surfaces. So far, very little has been revealed about the mobility of their phycobilisomes and the regulation of their light-harvesting system in general. Therefore, we carried out a detailed analysis of phycobilisome dynamics in several red alga strains and compared these results with the presence (or absence) of photoprotective mechanisms. Our data conclusively prove phycobilisome mobility in two model mesophilic red alga strains, Porphyridium cruentum and Rhodella violacea. In contrast, there was almost no phycobilisome mobility in the thermophilic red alga Cyanidium caldarium that was not caused by a decrease in lipid desaturation in this extremophile. Experimental data attributed this immobility to the strong phycobilisome-photosystem interaction that highly restricted phycobilisome movement. Variations in phycobilisome mobility reflect the different ways in which light-harvesting antennae can be regulated in mesophilic and thermophilic red algae. Fluorescence changes attributed in cyanobacteria to state transitions were observed only in mesophilic P. cruentum with mobile phycobilisomes, and they were absent in the extremophilic C. caldarium with immobile phycobilisomes. We suggest that state transitions have an important regulatory function in mesophilic red algae; however, in thermophilic red algae, this process is replaced by nonphotochemical quenching.Photosynthetic light reactions are mediated by pigment-binding protein complexes located either inside the thylakoid membrane (e.g. chlorophyll-binding proteins of both photosystems) or associated on the membrane surface (e.g. phycobilisomes [PBsomes] in cyanobacteria and red algae). Recent progress in structural biology has allowed the construction of high-resolution structural models of most photosynthetic protein complexes (for review, see Fromme, 2008) together with their large-scale organization into supercomplexes (for review, see Dekker and Boekema, 2005). However, the dynamics of these supercomplexes and the mobility of particular light-harvesting proteins in vivo are still poorly understood (for review, see Mullineaux, 2008a; Kaňa, 2013; Kirchhoff, 2014) The importance of protein mobility in various photosynthetic processes, like nonphotochemical quenching and state transitions, has been explored mostly based on indirect in vitro experiments, including single-particle analysis (Kouřil et al., 2005), or by biochemical methods (Betterle et al., 2009; Caffarri et al., 2009). Recent studies on the mobility of light-harvesting proteins using live-cell imaging (for review, see Mullineaux, 2008a; Kaňa, 2013) have elucidated the importance of protein mobility for photosynthetic function (Joshua and Mullineaux, 2004; Joshua et al., 2005; Goral et al., 2010, 2012; Johnson et al., 2011). In addition, the redistribution of respiratory complexes in cyanobacterial thylakoid membranes plays an essential role in controlling electron flow (Liu et al., 2012).It is generally accepted that the mobility of most of the transmembrane photosynthetic proteins is very restricted in the thylakoid. The typical effective diffusion coefficient of photosynthetic proteins is somewhere between 0.01 and 0.001 μm⁻² s⁻¹ (Kaňa, 2013). A similar restriction in membrane protein mobility has also been described for bacterial membranes (Dix and Verkman, 2008; Mika and Poolman, 2011). In fact, this is very different in comparison with what we know for other eukaryotic membranes (e.g. plasma membrane and endoplasmic reticulum), where membrane-protein diffusion can be faster by 1 or 2 orders of magnitude (Lippincott-Schwartz et al., 2001). Therefore, macromolecular crowding of proteins has been used to rationalize the restricted protein mobility in thylakoid membranes of chloroplasts (Kirchhoff, 2008a, 2008b). Indeed, atomic force microscopy studies have shown that there is a dense packing and interaction of complexes in the photosynthetic membranes (Liu et al., 2011). Therefore, the diffusion of photosynthetic proteins in the thylakoid membrane is rather slow, and it increases only in less crowded parts of thylakoids (Kirchhoff et al., 2013). The current model of photosynthetic protein mobility thus proposes the immobility of protein supercomplexes, such as PSII (Mullineaux et al., 1997; Kirchhoff, 2008b), with only a small mobile fraction of chlorophyll-binding proteins represented by external antennae of photosystems, including light harvesting complex of PSII in higher plants (Consoli et al., 2005; Kirchhoff et al., 2008) or iron stress-induced chlorophyll-binding protein A in cyanobacteria (Sarcina and Mullineaux, 2004).The restricted mobility of internal membrane supercomplexes (photosystems) contrasts with the relatively mobile PBsomes (Mullineaux et al., 1997; Sarcina et al., 2001). PBsomes are sizeable biliprotein supercomplexes (5–10 MD) attached to the thylakoid membrane surface with dimensions of approximately 64 × 42 × 28 nm (length × width × height; Arteni et al., 2008; Liu et al., 2008a). PBsomes are composed of chromophore-bearing phycobiliproteins and colorless linker polypeptides (Adir, 2005; Liu et al., 2005). They serve as the main light-harvesting antennae in various species, including cyanobacteria, red algae, glaucocystophytes, and cryptophytes. Although a single PBsome is composed of hundreds of biliproteins, absorbed light energy is efficiently transferred toward a specific biliprotein that functions as a terminal energy emitter (Glazer, 1989). From there, energy can be transferred to either PSI or PSII and used in photosynthesis (Mullineaux et al., 1990; Mullineaux, 1992, 1994). In typical prokaryotic cyanobacteria and eukaryotic red algae, PBsomes are composed of two main parts: (1) allophycocyanin (APC) core proteins adjacent to the thylakoid membrane; and (2) peripheral rod proteins made from phycocyanin only or from a combination of phycocyanin together with phycoerythrin. Such complex and modular composition allows for different spectroscopic properties of PBsomes and thus their complementary absorption in the spectral region that is not covered by chlorophyll-binding proteins.PBsome mobility has been studied only in a few types of cyanobacteria (for review, see Kaňa, 2013). PBsomes have been recognized as a mobile element with an effective diffusion coefficient of about 0.03 μm² s⁻¹ for Synechococcus sp. PCC 7942 (Mullineaux et al., 1997; Sarcina et al., 2001). The effective diffusion coefficient value depends on lipid composition, temperature, and the size of the PBsome (Sarcina et al., 2001). The diffusion coefficient reflects PBsome mobility, but it is not affected singularly by physical diffusion processes, and the role of PBsome-photosystem interaction is an open question (Kaňa, 2013). PBsome mobility seems to be related to the requirement of light-induced PBsome redistribution during state transitions (Joshua and Mullineaux, 2004). The mechanism of state transitions in cyanobacteria is still rather questionable (for review, see Kirilovsky et al., 2014). As PSII seems to be immobile, it has been suggested that PBsomes interact with photosystems only transiently and that physical redistribution (diffusion) of PBsomes is crucial for the state transition (Mullineaux et al., 1997). The importance of such long-distance diffusions, however, should be tested experimentally in more detail (Kaňa, 2013), as an alternative theory of the state transition proposed only slight PBsome movement (shifting) between photosystems (McConnell et al., 2002). However, in both cases, PBsome mobility (i.e. the PBsome’s ability to move) is required (Kaňa, 2013).Red algae are the eukaryotic representatives of phototrophs containing PBsomes (Su et al., 2010). They represent the ancestor of photosynthetic microorganisms from the red clade of photosynthesis (Yoon et al., 2006; Wang et al., 2013), which includes various model organisms such as diatoms, chromerids, or dinoflagellates. Red algae contain a unique combination of antennae systems on their membrane surfaces, which are formed mostly by hemispherical PBsomes (Mimuro and Kikuchi, 2003; Arteni et al., 2008). Red algae also contain transmembrane light-harvesting antennae (Vanselow et al., 2009; Neilson and Durnford, 2010; Green, 2011) associated mostly with PSI (Wolfe et al., 1994). Therefore, red algae represent a functionally important eukaryotic model organism; however, few facts are known about the regulation of its light-harvesting efficiency, although it seems to be connected with photoprotection in the reaction center (Delphin et al., 1996, 1998; Krupnik et al., 2013). The presence of photoprotective NPQ in PBsomes of prokaryotic cyanobacteria has been conclusively proven (Kirilovsky et al., 2014); however, this mechanism seems to be missing in eukaryotic phycobiliproteins of cryptophytes (Kaňa et al., 2012b) and red algae. Moreover, the presence (or absence) of PBsome mobility has not been confirmed conclusively (Liu et al., 2009).Therefore, we carried out a detailed study of PBsome mobility in red algal chloroplasts to determine the role of mobility in the regulation of light-harvesting efficiency. We found that red alga PBsomes are a mobile protein complex with effective diffusion coefficient between 2.7 × 10⁻³ and 13 × 10⁻³ μm⁻² s⁻¹ in all studied mesophilic strains. It contrasted with PBsomes in extremophilic red algal strains (Cyanidium caldarium), where PBsome mobility under physiological conditions was highly restricted (effective diffusion coefficient of approximately 0.6 × 10⁻³ μm⁻² s⁻¹). The restriction of PBsome mobility in extremophilic C. caldarium was due to a tight interaction of PBsomes with both photosystems and not to changes in lipid desaturation, an effect typical for extremophiles. The PBsome-photosystem interaction was weakened for C. caldarium grown at suboptimal temperatures, resulting in a pronounced increase in PBsome mobility thanks to PBsome decoupling from the photosystem. This result shows that PBsome mobility in this strain is limited by the strength of the PBsome-photosystem interaction rather than by the restriction of diffusion by factors such as macromolecular crowding. Moreover, our study allows us to describe two different models of light-harvesting antenna regulation in red algae. In mesophilic strains (Porphyridium cruentum and Rhodella violacea), absorbed light is redistributed between photosystems in a process of state transition that requires PBsome mobility. On the contrary, in extremophilic C. caldarium, PBsome are strongly coupled to photosystems and excess light is dissipated by a process of nonphotochemical quenching, as has been described recently (Krupnik et al., 2013).     

念珠藻生长特性研究

研究了念珠藻的生长周期、固氮特性、生长条件等生长特性.得出本实验念珠藻可以固氮;磷能有效制约其生长;需要长时间的光照等结果.     

Phycobilisome structure in the cyanobacteria Mastigocladus laminosus and Anabaena sp. PCC 7120.

Phycobilisomes of the cyanobacteria Mastigocladus laminosus and Anabaena sp. PCC7120 differ from typical tricylindrical, hemidiscoidal phycobilisomes in three respects. Firstly, size comparisons of the core-membrane linker phycobiliproteins (LCM) in different cyanobacteria by SDS/PAGE reveal an apparent molecular mass of 120 kDa for the LCM of M. laminosus and Anabaena sp. PCC7120. This observation suggests that the polypeptides of these species have four linker-repeat domains. Secondly, phycobilisomes of M. laminosus are shown to contain at least three, but most probably four, different rod-core linker polypeptides (LRC). These LRC, which attach the peripheral rods to the core and thereby make phycocyanin/allophycocyanin contacts, have been identified and characterized by N-terminal amino acid sequence analysis. Additionally, electron microscopy of phycobilisomes isolated from M. laminosus and Anabaena sp. PCC7120 reveals similar structures which differ from those of Calothrix sp. PCC7601 with their typical six, peripheral rods. Based upon protein-analytical results and a reinterpretation of the data of [Isono, T. & Katoh, T. (1987) Arch. Biochem. Biophys. 256, 317-324], we discuss structural implications of recent findings on the established hemidiscoidal model for the phycobilisomes of M. laminosus and Anabaena sp. PCC7120. Up to eight peripheral rods are suggested to radiate from a modified core substructure which contains two additional peripheral allophycocyanin hexamer equivalents that serve as the core-proximal discs for two peripheral rods.     

Phycobilisome Mobility in the Cyanobacterium Synechococcus sp. PCC7942 is Influenced by the Trimerisation of Photosystem I

We have constructed a mutant of the cyanobacterium Synechococcus sp. PCC7942 deficient in the Photosystem I subunit PsaL. As has been shown in other cyanobacteria, we find that Photosystem I is exclusively monomeric in the PsaL(-) mutant: no Photosystem I trimers can be isolated. The mutation does not significantly alter pigment composition, photosystem stoichiometry, or the steady-state light-harvesting properties of the cells. In agreement with a study in Synechococcus sp. PCC7002 [Schluchter et al. (1996) Photochem Photobiol 64: 53-66], we find that state transitions, a physiological adaptation of light-harvesting function, occur significantly faster in the PsaL(-) mutant than in the wild-type. To explore the reasons for this, we have used fluorescence recovery after photobleaching (FRAP) to measure the diffusion of phycobilisomes in vivo. We find that phycobilisomes diffuse, on average, nearly three times faster in the PsaL(-) mutant than in the wild-type. We discuss the implications for the mechanism of state transitions in cyanobacteria.     

Identification of sesquiterpene synthases from Nostoc punctiforme PCC 73102 and Nostoc sp. strain PCC 7120

Cyanobacteria are a rich source of natural products and are known to produce terpenoids. These bacteria are the major source of the musty-smelling terpenes geosmin and 2-methylisoborneol, which are found in many natural water supplies; however, no terpene synthases have been characterized from these organisms to date. Here, we describe the characterization of three sesquiterpene synthases identified in Nostoc sp. strain PCC 7120 (terpene synthase NS1) and Nostoc punctiforme PCC 73102 (terpene synthases NP1 and NP2). The second terpene synthase in N. punctiforme (NP2) is homologous to fusion-type sesquiterpene synthases from Streptomyces spp. shown to produce geosmin via an intermediate germacradienol. The enzymes were functionally expressed in Escherichia coli, and their terpene products were structurally identified as germacrene A (from NS1), the eudesmadiene 8a-epi-α-selinene (from NP1), and germacradienol (from NP2). The product of NP1, 8a-epi-α-selinene, so far has been isolated only from termites, in which it functions as a defense compound. Terpene synthases NP1 and NS1 are part of an apparent minicluster that includes a P450 and a putative hybrid two-component protein located downstream of the terpene synthases. Coexpression of P450 genes with their adjacent located terpene synthase genes in E. coli demonstrates that the P450 from Nostoc sp. can be functionally expressed in E. coli when coexpressed with a ferredoxin gene and a ferredoxin reductase gene from Nostoc and that the enzyme oxygenates the NS1 terpene product germacrene A. This represents to the best of our knowledge the first example of functional expression of a cyanobacterial P450 in E. coli.     

Reduction of Photoautotrophic Productivity in the Cyanobacterium Synechocystis sp. Strain PCC 6803 by Phycobilisome Antenna Truncation

Truncation of the algal light-harvesting antenna is expected to enhance photosynthetic productivity. The wild type and three mutant strains of Synechocystis sp. strain 6803 with a progressively smaller phycobilisome antenna were examined under different light and CO(2) conditions. Surprisingly, such antenna truncation resulted in decreased whole-culture productivity for this cyanobacterium.     

Characterization of the glutamate/aspartate-transport system in a symbiotic Nostoc sp.

The permeability properties of the cell membrane of a symbiotic Nostoc sp. for glutamate and aspartate were investigated. These compounds were translocated across the plasmalemma by a transport system which showed a very high affinity for glutamate and a lower one for aspartate. Since a concomitant release of glutamate was observed during the uptake of these two amino acids it is concluded that the transport proceeds via a counterexchange mechanism. In addition to this counterexchange a net release of glutamate occurred in the dark. Some aspects concerning the possible function of this transport system in the symbiotic association Geosiphon pyriforme are discussed.     

Freeze-Recovery Physiology of Nitrogenase Activity in Terrestrial Nostoc sp. Colonies

Nostoc sp. colonies from field collections were cultured and propagated on silica sand with aqueous N-free BG-11 medium. Laboratory experiments were conducted to characterize the in vivo freeze-recovery physiology of nitrogenase activity. Nitrogenase activity was monitored by the acetylene reduction technique. Frozen Nostoc sp. colonies were thawed and warmed to 10, 15, 20, 25, or 30°C. At 25 and 30°C, nitrogenase activity was detected within 6 h after thawing. At 20°C or lower, nitrogenase activity was not detected until 12 h after thawing. Optimum thawing temperature with respect to the recovery of nitrogenase activity was 25°C. In subsequent experiments, laboratory-grown Nostoc colonies were used along with the following conditions: prefreezing treatment of 3 days of exposure to light or darkness, freezing, and then thawing to 25°C in light or darkness with or without metabolic inhibitors [3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU), monofluoroacetate, or chloramphenicol]. Approximately 30% of the energy in the initial recovery of nitrogenase activity (to 12 h after thawing) appeared to be supplied via the utilization of carbon compounds stored before freezing. Photosynthetic conditions (i.e., light and without DCMU) were necessary for maximum recovery of nitrogenase activity. In the presence of the protein synthesis inhibitor chloramphenicol, nitrogenase activity was still detected at 12 to 48 h after thawing. Although damage may occur to nitrogenase, some of the enzyme was capable of surviving the freeze-thaw period in vivo. However, complete recovery of nitrogenase activity (equal to prefreezing activity) may entail some de novo synthesis of nitrogenase.     

Relationship between color and pigment production in two stone biofilm-forming cyanobacteria (Nostoc sp. PCC 9104 and Nostoc sp. PCC 9025)

Previous studies have provided evidence that color measurements enable on site quantification of superficial biofilms, thereby avoiding the need for sampling. In the present study, the efficiency of color measurements to evaluate to what extent pigment production is affected by environmental parameters such as light intensity, combined nitrogen and nutrient availability, was tested with two cyanobacteria, Nostoc sp. strains PCC 9104 and PCC 9025, which form biofilms on stone. Both strains were acclimated, in aerated batch cultures for 2 weeks, to three different culture media: BG-11, BG-11₀, and BG-11₀/10 at either high or low light intensity. The content of chlorophyll a, carotenoids, and phycocyanins was measured throughout the experiment, together with variations in the color of the cyanobacteria, which were represented in the CIELAB color space. The results confirmed that the CIELAB color parameters are correlated with pigment content in such a way that variations in the latter are reflected as variations in color.     

Diversity and transcription of proteases involved in the maturation of hydrogenases in Nostoc punctiforme ATCC 29133 and Nostoc sp. strain PCC 7120

Background  

The last step in the maturation process of the large subunit of [NiFe]-hydrogenases is a proteolytic cleavage of the C-terminal by a hydrogenase specific protease. Contrary to other accessory proteins these hydrogenase proteases are believed to be specific whereby one type of hydrogenases specific protease only cleaves one type of hydrogenase. In cyanobacteria this is achieved by the gene product of either hupW or hoxW, specific for the uptake or the bidirectional hydrogenase respectively. The filamentous cyanobacteria Nostoc punctiforme ATCC 29133 and Nostoc sp strain PCC 7120 may contain a single uptake hydrogenase or both an uptake and a bidirectional hydrogenase respectively.     

Nutritional evaluation of Cyanobacterium (Nostoc sp.) extract in Rhizobium cultures

Extracts from Nostoc sp. biomass (CE) were compared with yeast extract (YE) in the formulation of media for cultivation of Rhizobium etli and Bradyrhizobium japonicum. Growth depended on the rhizobium strain and also on the type and concentration of the extract. The growth kinetics of R. etli were substantially influenced by the addition of CE an YE at concentrations between 0.3 and 1% (w/v); cultures containing YE showed shorter lag periods and higher growth rates and those with CE presented higher growth yields. Depending on the extracts, a different behaviour was displayed by the cultures at the stationary phase; cell viability increased or remained fairly constant in cultures developed on CE, and those developed on high concentrations of YE showed a decline in viable cell counts.     

Phycobilisome Heterogeneity in the Red Alga Porphyra umbilicalis

Phycobilisomes were isolated from Rhodophyceae brought from the field (Porphyra umbilicalis) or grown in culture under laboratory conditions (Antithamnion glanduliferum). In P. umbilicalis two kinds of well-coupled (ellipsoidal and hemidiscoidal) phycobilisomes were detected, in contrast to A. glanduliferum cultured algae in which only one kind of well-coupled, ellipsoidaltype phycobilisome appeared. The new phycobilisome-type particle detected in P. umbilicalis is characterized by an impoverishment in R-phycoerythrin and by sedimentation at lower density. The comparison between both phycobilisomes of P. umbilicalis allows determination of the presence of one colorless linker polypeptide (30 kilodaltons) associated with R-phycocyanin and allophycocyanin and two (40 and 38 kilodaltons) associated to R-phycoerythrin. The percentage of linker polypeptides associated with this pigment is low in the new phycobilisome-like particle detected. This suggests that part of the R-phycoerythrin is less strongly bound to the phycobilisome than the other pigments. This feature could probably explain the existence of two kinds of phycobilisomes as intermediary steps of phycobilisome organization in algae exposed to rapid changes in environmental factors. In contrast, algae growing in culture and adapted to specific conditions do not present intermediary organization steps. Polypeptide composition and identification are given for this phycobilisome-like particle.     

Synthesis and Assembly of the Polypeptides of Photosystem I and II in Isolated Etiochloroplasts of Wheat

Synthesis and assembly of photosystems (PS) I and II polypeptides in etiochloroplasts isolated from greening wheat (Triticum aestivum L. cv Norin 61) seedlings were studied. The isolated etiochloroplasts synthesized PSI polypeptides of 66 and 15 kilodaltons, PSII polypeptides of 46 and 42 kilodaltons, and atrazine-binding 34 to 32 kilodalton polypeptide. Their assembly processes in the thylakoid membrane were studied by pulse-chase labeling with [³⁵S]methionine, mild solubilization of the thylakoid membrane with Triton X-100, sucrose density gradient centrifugation, and polyacrylamide gel electrophoresis. The newly synthesized polypeptides of 66, 46, 42, 34, and 32 kilodaltons were first integrated into the complexes of 7.5, 5.9, 7.5, 6.3, and 7.5 Svedberg units, respectively, in 20 minutes. After the chase with excess amount of methionine for 100 min, they were found in complexes of 9.5, 9.1, 9.1, 9.1, and 9.1 Svedberg units, respectively. In this condition, stained polypeptides of PSI and PSII were found in the complexes of 11.1 and 10.3 Svedberg units, respectively. These results indicated that newly synthesized PSI or PSII polypeptides are integrated into intermediate complexes, but not complete complexes in the isolated etiochloroplasts. The relationship between the processing of the atrazine-binding 32 kilodalton polypeptide and its assembly into the PSII complex is also discussed.