Activation of sulphate in Anabaena cylindrica

Summary Crude cell-free extracts of Anabaena cylindrica synthesized adenosine-5-phosphosulphate (AP³⁵S) and 3-phosphoadenosine-5-phosphosulphate (PAP³⁵S) from ³⁵SO₄ ^2- in the presence of Mg²⁺, ATP and inorganic pyrophosphatase. Maximum AP³⁵S and PAP³⁵S were produced at pH 7.15 and 8.05, respectively. APS kinase was detected in the supernatant of crude cell-free extracts by a spectrophotometric procedure. ATP-Sulphurylase had an absolute requirement for Mg²⁺ and less than 30% AP³⁵S was formed when Mg²⁺ was replaced by either Mn²⁺ or Co²⁺. Nucleotide triphosphates other than ATP and 2-deoxyATP were ineffective in this reaction. Maximum enzyme activity was observed at equimolar concentrations of Mg²⁺ and ATP and excess of either of these was inhibitory. Other nucleotide triphosphates, like GTP, UTP, CTP, TTP, ITP, or 2-deoxyATP also inhibited the enzyme activity. Inhibition by GTP was competitive with respect to ATP. ATP-sulphurylase activity was not affected by cysteine, methionine or glutathione.Abbreviations APS adenosine-5-phosphosulphate - PAPS 3-phosphoadenosine-5-phosphosulphate     

Nitrogen fixation by Anabaena cylindrica

Summary Blending Anabaena cylindrica cultures results in a loss of nitrogenase activity which is correlated with the breakage of the filaments at the junctions between heterocysts and vegetative cells. Oxygen inhibition of nitrogen fixation was significant only above atmospheric concentrations. Nitrogen-fixation activities in the dark were up to 50% of those observed in the light and were dependent on oxygen (10 to 20% was optimal). Nitrogenase activity was lost in about 3 h when cells were incubated aerobically in the dark. Re-exposure to light resulted in recovery of nitrogenase activity within 2 h. Blending, oxygen, or dark pre-incubation had similar effects upon cultures grown under air or nitrogen and did not inhibit light-dependent CO₂ fixation. We conclude that heterocysts are the sites of nitrogenase activity and propose a model for nitrogen fixation by Anabaena cylindrica.     

Nitrogen fixation by Anabaena cylindrica

Summary The effects of oxygen, light and photosynthesis inhibitors on nitrogenase activities in Anabaena cylindrica batch cultures were followed as a function of time after inoculation. During the early rapid growth period the nitrogenase activities of cultures grown under air/CO₂ or N₂/CO₂ were relatively resistant to oxygen and DCMU inhibition. These cultures also exhibited oxygen-dependent nitrogenase activity in the dark of up to 50% of that measured in the light. After active growth ceased the cultures continued to slowly grow for a prolonged period of time. The nitrogenase activities of these old cultures were very sensitive to oxygen and DCMU inhibition. These cultures also had little or no dark nitrogenase activities. The photosynthesis inhibitor DBMIB was not a specific inhibitor of light-driven electron transport since it inhibited both light and dark nitrogenase activities. Nitrogenase activities induced under oxygen-free/CO₂ gas mixtures initially were significantly more sensitive to oxygen inhibition than those induced under air/CO₂. We discuss these results in relation to heterocyst function.     

The fatty acid composition of akinetes, heterocysts and vegetative cells in Anabaena cylindrica

The fatty acid composition of akinetes, heterocysts and vegetativecells in Anabaena cylindrica was examined. Akinetes and heterocystscontained much less -linolenic acid than did vegetative cells.Furthermore, akinetes and heterocysts contained fatty acidswith less unsaturation as compared with vegetative cells. (Received February 19, 1972; )     

Nitrogen fixation by Anabaena cylindrica

Hydrogen-supported nitrogenase activity was demonstrated in Anabaena cylindrica cultures limited for reductant. Nitrogen-fixing Anabaena cylindrica cultures sparged in the light with anaerobic gases in the presence of the photosynthesis inhibitor DCMU slowly lost their ability to reduce acetylene in the light under argon but exhibited near normal activities in the presence of 11% H₂ (balance argon). The hydrogen-supported nitrogenase activity was half-saturated between 2 and 3% H₂ and was strongly inhibited by oxygen (50% inhibition at about 5–6% O₂). Batch cultures of Anabaena cylindrica approaching stationary growth phase (“old” cultures) lost nitrogenase-dependent hydrogen evolution almost completely. In these old cultures hydrogen relieved the inhibitory effects of DCMU and O₂ on acetylene reduction. Our results suggest that heterocysts contain an uptake hydrogenase which supplies an electron transport chain to nitrogenase but which couples only poorly with the respiratory chain in heterocysts and does not function in CO₂ fixation by vegetative cells.     

Protective mechanisms in photosynthesis of Anabaena cylindrica

The role of O₂ photoreduction was studied in intact cells of normal and photobleached Anabaena cylindrica Lemm. strain PCC 7122. We found that O₂ photoreduction represents a protective mechanism against over-reduction of the photosyn-thetic electron transport chain only in normal Anabaena cells. This protective mechanism was not functioning in photobleached cells in spite of the increased rate of photosynthetic electron flow. A new electron acceptor, the induced reversible hydrogenase, is suggested to be operating in photobleached Anabaena cylindrica .     

Hydrogen evolution by photobleached Anabaena cylindrica

We have studied the evolution of hydrogen by photobleached filaments of the heterocystous bluegreen alga Anabaena cylindrica. The photobleached cells became orange-yellow due to the heavy accumulation of carotenoids. We found that the yellow filaments produced much larger amounts of hydrogen than the normal, green ones, while the nitrogenase activity responsible for hydrogen evolution increased to a lesser extent. We suggest that a reversible hydrogenase activity induced in photobleached filaments is responsible for the excess amount of hydrogen. 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU) inhibits the hydrogen evolution of the yellow filaments which produce much more oxygen and fix less CO₂ than the green filaments. Therefore we consider the water to be a possible electron source for this hydrogenase. The low efficiency of light energy conversion (0.3%) in nitrogenase-catalyzed H₂ evolution (Laczkó, 1980 Z. Pflanzenphysiol. 100, 241–245) is increased to 1.5–2% by the appearance of the reversible hydrogenase activity.Abbreviations Chl chlorophyll - Car carotenoids - Phy phycocyanin - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl-urea - PSI photosystem I - PSII photosystem II     

GTP-binding proteins in a cyanobacterium Anabaena cylindrica

GTP-binding proteins were detected in a crude extract containing membrane components of Anabaena cylindrica. The crude extract was treated with 1% Lubrol PX and was fractionated by gel filtration. The binding of [35S]GTP gamma S to GTP-binding proteins was prevented in the presence of 0.1 mM GTP and in the presence of 0.1 mM ATP. Six fractions of these GTP-binding proteins, tentatively designated GA1 to GA6, were ADP-ribosylated by pertussis toxin. GA3, GA4 and GA5 had Km values of 10, 60 and 7 nM, respectively. The molecular weights of some of these GTP-binding proteins were reduced after being labelled with [35S]GTP gamma S.     

Sporulation in the filamentous cyanobacterium Anabaena cylindrica

Sporulation in the filamentous cyanobacterium Anabaena cylindrica involves the transformation of a vegetative cell into a thick-walled resistant structure. Because this process occurs at predictable loci in each filament and involves a significant increase in cell size, the course of sporulation in a culture can be quantitatively determined. Sporulation occurs during the late logarithmic phase of a culture, a time of slow but unbalanced growth. Under the conditions imployed here, sporulation is not a synchronous event either between or within filaments. The information in this paper provides an estimate of the rate of spore differentiation and supports the previous notion that in the formation of strings of more than one spore, a gradient of spore maturation exists.     

Light-induced accumulation of lactate and succinate in Anabaena cylindrica

In the cyanobacterium Anabaena cylindrica lactate accumulated in large amounts when the cells were exposed to light. The presence or absence of oxygen, or a change in CO₂ concentration did not affect the lactate accumulation. The cellular succinate level also increased in the light when CO₂ was supplied at the high concentration of 1%. 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU), an inhibitor of photosynthetic electron flow, inhibited the increase in the concentration of lactate and succinate. Photosynthesis is a prerequisite for the increase of these organic acids. Thenoyltrifluoroacetone, an inhibitor of succinate dehydrogenase, inhibited the increase of succinate, suggesting that the succinate is formed via fumarate by the reverse of reactions of tricarboxylic acid (TCA) cycle. Upon addition of ammonium to the cell suspension in the light under high CO₂ concentration, the increases in the concentrations of lactate and succinate were inhibited while those of glutamine, glutamate and aspartate were stimulated. Ammonium apparently changed the products of metabolism of pyruvate and oxaloacetate from lactate and succinate to amino acids.Abbreviations Chl chlorophyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - TTFA thenoyltrifluoroacetone - PCA perchloric acid     

Induction of nitrate and nitrite reductases in Anabaena cylindrica

Induction of nitrate and nitrite reductases in Anabaena cylindrica(FOGG strain) was investigated. At various stages of algal growthin the presence of nitrate or nitrite, the levels of these enzymeswere determined using cell-free preparations. Nitrate and nitritereductases were induced by the respective substrates. Nitratedid not act either as an inducer or as a repressor of nitritereductase. ¹This work was supported by grant No. 8814 from the Ministryof Education ²Department of Biology, Faculty of Science, Tokyo MetropolitanUniversitySetagaya-ku, Tokyo, Japan (Received June 18, 1970; )     

盐胁迫对柱胞鱼腥藻细胞结构和固氮作用的影响

在含KCl的条件下培养,柱胞鱼腥藻细胞膨大,球形化,色素质靠向细胞一侧,另一侧变成无色透明区。电镜检查,无色透明区形成液泡。此种结构改变是可逆的,CaCl2可抵消KCl的这种作用。在含KCl的无氮培养基中培养,柱胞鱼腥藻生长迟缓,细胞黄化,乙炔还原法测定固氮活性下降95％。     

Mutants of Anabaena cylindrica altered in heterocyst spacing

Nitrosoguanidine induced mutants of Anabaena cylindrica have been obtained, which are altered in heterocyst spacing. In the wild type organism the pattern is composed of single intercalary heterocysts. The mutant patterns fall into several classes: those with only terminal heterocysts, with both terminal and intercalary heterocysts, with groups of heterocysts and those totally lacking heterocysts. The mutants are described in detail, and the various pattern modifications are interpreted in terms of a model we have proposed.     

Changes in thylakoid structure associated with the differentiation of heterocysts in the cyanobacterium, Anabaena cylindrica.

The thylakoids of vegetative cells of the filamentous cyanobacterium, Anabaena cylindrica, are capable of oxygen-evolving photosynthesis and contain both Photosystems I and II (PSI and PSII). The heterocysts, cells specialized for nitrogen fixation, do not produce oxygen and lack Photosystem II activity, the major accessory pigments, and perhaps the chlorophyll a associated with PSII. Freeze-fracture replicas of vegetative cells and of heterocysts reveal differences in the structure of the thylakoids. A histogram of particle sizes on the exoplasmic fracture face (E-face, EF) of vegetative cell thylakoids has two major peaks, at 75 and 100 A. The corresponding histogram for heterocyst thylakoids lacks the 100 A size class, but has a very large peak at about 55 A with a shoulder at 75 A. Histograms of protoplasmic fracture face (P-face, PF) particle diameters show single broad peaks, the mean diameter being 71 A for vegetative cells and 64 A for heterocysts. The thylakoids of both cell types have about 5600 particles/micrometers2 on the P-face. On the E-face, the density drops from 939 particles/micrometers2 on vegetative cell thylakoids to 715 particles/micrometers2 on heterocyst thylakoids. The data suggest that the 100 A E-face particle of vegetative cell thylakoids is a PSII complex. The 55 A EF particle of heterocysts may be part of the nitrogenase complex or a remnant of the PSII complex. The role of the 75 A EF particle is unknown. Other functions localized on cyanobacterial thylakoids, such as respiration and hydrogenase activity, must be considered when interpreting the structure of these complex thylakoids.     

Metabolic Activities of Isolated Spores of Anabaena cylindrica

A method for the isolation of spores of Anabaena cylindricais described, and results concerning the metabolic activitiesof isolated spores are presented. Isolated spores fixed carbondioxide in the light at a lower rate and evolved carbon dioxidein the dark at a higher rate than the corresponding whole filaments.Nitrogen-fixing ability in isolated spores could be detectedneither with the isotopic method nor with the new acetylenetechnique. Isolated spores were able to germinate at the samerate as spores within the original cell population when transferredinto fresh culture medium.