Primary photosynthetic processes in Heliobacterium chlorum at 15K

Absorbance changes induced by 25-ps laser flashes were measured in membranes of Heliobacterium chlorum at 15 K. Absorbance difference spectra, measured at various times after the flash showed negative bands in the Q_y region at 812, 793 and 665 nm. The first of these bands was attributed to the formation of excited singlet states of a long-wavelength form of antenna bacteriochlorophyll g (BChl g 808). Absorbance changes of shorter wavelength absorbing antenna BChls g were at least an order of magnitude smaller, indicating rapid excitation energy transfer (i.e. within the time resolution of the apparatus) from these BChls to BChl g 808. Excited BChl g 808 showed a bi-exponential decay with time constants of 50 and 200 ps. The bands at 793 and 665 nm may be attributed to the primary charge separation and reflect the photooxidation of the primary electron donor P-798 and photoreduction of a primary electron acceptor absorbing near 670 nm, presumably a BChl c or Chl a-like pigment. The bleaching of this pigment reversed with a time constant of 300 ps at 15 K and of 800 ps at 300 K. This indicates that electron transfer from the primary to the secondary electron acceptor is approximately 2.5 times faster at 15 K than at room temperature.Abbreviations BChl bacteriochlorophyll - FWHM full width at half maximum - P-798 primary electron donor - Tris tris(hydroxymethyl)amino methane     

Single-stranded DNA used as an efficient new vehicle for transformation of plant protoplasts

In relation to the question which DNA form (single- or double-stranded) is transferred by Agrobacterium tumefaciens to plant cells, we studied the behaviour of single-stranded DNA, as compared to double-stranded DNA, when it is introduced into plant protoplasts by electroporation. To this end, we cloned a construct with a plant NPTII gene as well as a CAT gene in the M13 vectors tg130 and tg131. We found that both complementary single-stranded molecules gave rise to substantial CAT activity in plant protoplasts, suggesting that single-stranded DNA is converted into double-stranded DNA by the plant cell replication machinery. Unexpectedly, we found that single-stranded DNA leads to a 3–10 fold higher frequency of stable transformation (selection for kanamycin resistance) than double-stranded DNA. These results indicate that the use of single-stranded DNA might be considered in experiments in which optimal transformation frequencies are needed, e.g. with protoplasts form recalcitrant plant species.Abbreviations ss single-stranded - ds double-stranded - CAT chloramphenicol acetyl transferase - NPTII neomycin phosphotransferase II - RT room temperature     

Transformation of Claviceps purpurea using a bleomycin resistance gene

Summary To develop a DNA-mediated transformation system for Claviceps purpurea a vector was constructed using a bleomycin-resistance gene (bleo ^R) fused in frame to the Aspergillus nidulans trp C promoter as a dominant selection marker. The construct was shown to be functional in Aspergillus nidulans and Aspergillus niger and used to transform a wild strain of Claviceps purpurea. Transformats were obtained at low frequencies; they were shown to contain transforming DNA integrated into the chromosomal DNA, probably in multimeric copies and at multiple sites. Combined Southern, Northern and resistance level analysis indicate that the A. nidulans promoter is functional in C. purpurea.Dedicated to Professor Dr. Dr. h. c. K. Esser on the occasion of his 65^th birthday     

The influence of experimental conditions on the spectrin-hemoglobin interaction

Human spectrin, when isolated, purified and stored in such conditions that preserve its tetrameric form, is able to associate with human hemoglobin as it is clearly shown by gel filtration. However, this hemoglobin-spectrin association does not seem to have a significant effect on hemoglobin oxygenation as indicated by equilibrium and rapid kinetics measurements.     

Immunocytochemical and ultrastructural study of the frog (Rana pipiens) pars distalis with special reference to folliculo-stellate cell function during in vitro superfusion

Summary The colloidal gold immunocytochemical technique was used to determine the ultrastructural features of the glandular cells in the pituitaries of male frogs, Rana pipiens, both in vivo and after superfusion in vitro. Specific reactions to antisera against bullfrog gonadotropins, human prolactin, and synthetic 1–39 corticotropin allowed identification of the 3 corresponding types of glandular cells. No immunoreaction was obtained with antisera against human or ovine-growth hormone, human -thyrotropin hormone, and bovine S-100 protein. General morphological features of these immunocytochemically identified glandular cells were similar to those of equivalent cells previously described in other amphibian species. Non-glandular folliculo-stellate cells were distinctive. In freshly removed pituitaries, these folliculo-stellate cells contained lysosome-like structures, but did not show phagocytic vacuoles in the cytoplasm; they contained many mitochondria, and the Golgi complex and endoplasmic reticulum were relatively undeveloped. After 4 or 18 h of superfusion, some immunoreactive gonadotropic, prolactin, and corticotropic cells showed degeneration and destruction. In the same gland, folliculo-stellate cells retained a viable appearance, but showed phagocytic vacuoles containing secretory granule-like structures which were immunoreactive to gonadotropic, prolactin, and corticotropic antibodies. Some folliculo-stellate cells showed phagocytic vacuoles containing complete glandular cells. These results suggest that superfusion causes a destruction of some of the glandular cells, and that folliculo-stellate cells act as phagocytes when cellular debris or moribund cells are present in the intercellular space in the pituitary parenchyma.Supported by grant DCB 8710462 from the National Science Foundation, grant 2148-83 from the CAICYT (Spain) and the Junta de Andalucia (Spain)     

The role of subunit 4, a nuclear-encoded protein of the F0 sector of yeast mitochondrial ATP synthase, in the assembly of the whole complex

The yeast nuclear gene ATP4, encoding the ATP synthase subunit 4, was disrupted by insertion into the middle of it the selective marker URA3. Transformation of the Saccharomyces cerevisiae strain D273-10B/A/U produced a mutant unable to grow on glycerol medium. The ATP4 gene is unique since subunit 4 was not present in mutant mitochondria; the hypothetical truncated subunit 4 was never detected. ATPase was rendered oligomycin-insensitive and the F1 sector of this mutant appeared loosely bound to the membrane. Analysis of mitochondrially translated hydrophobic subunits of F0 revealed that subunits 8 and 9 were present, unlike subunit 6. This indicated a structural relationship between subunits 4 and 6 during biogenesis of F0. It therefore appears that subunit 4 (also called subunit b in beef heart and Escherichia coli ATP synthases) plays at least a structural role in the assembly of the whole complex. Disruption of the ATP4 gene also had a dramatic effect on the assembly of other mitochondrial complexes. Thus, the cytochrome oxidase activity of the mutant strain was about five times lower than that of the wild type. In addition, a high percentage of spontaneous rho- mutants was detected.     

Publisher's announcement

Publisher's announcement     

Specificity of Cellular DNA-Binding Sites of Microbial Populations in a Florida Reservoir

The substrate specificity of the DNA-binding mechanism(s) of bacteria in a Florida reservoir was investigated in short- and long-term uptake studies with radiolabeled DNA and unlabeled competitors. Thymine oligonucleotides ranging in size from 2 base pairs to 19 to 24 base pairs inhibited DNA binding in 20-min incubations by 43 to 77%. Deoxynucleoside monophosphates, thymidine, and thymine had little effect on short-term DNA binding, although several of these compounds inhibited the uptake of the radiolabel from DNA in 4-h incubations. Inorganic phosphate and glucose-1-phosphate inhibited neither short- nor long-term binding of [³H]- or [³²P]DNA, indicating that DNA was not utilized as a phosphorous source in this reservoir. RNA inhibited both short- and long-term radiolabeled DNA uptake as effectively as unlabeled DNA. Collectively these results indicate that aquatic bacteria possess a generalized nucleic acid uptake/binding mechanism specific for compounds containing phosphodiester bonds and capable of recognizing oligonucleotides as short as dinucleotides. This binding site is distinct from nucleoside-, nucleotide-, phosphomonoester-, and inorganic phosphate-binding sites. Such a nucleic acid-binding mechanism may have evolved for the utilization of extracellular DNA (and perhaps RNA), which is abundant in many marine and freshwater environments.     

Production of extracellular nucleic acids by genetically altered bacteria in aquatic-environment microcosms.

The factors which affect the production of extracellular DNA by genetically altered strains of Escherichia coli, Pseudomonas aeruginosa, Pseudomonas cepacia, and Bradyrhizobium japonicum in aquatic environments were investigated. Cellular nucleic acids were labeled in vivo by incubation with [3H]thymidine or [3H]adenine, and production of extracellular DNA in marine waters, artificial seawater, or minimal salts media was determined by detecting radiolabeled macromolecules in incubation filtrates. The presence or absence of the ambient microbial community had little effect on the production of extracellular DNA. Three of four organisms produced the greatest amounts of extracellular nucleic acids when incubated in low-salinity media (2% artificial seawater) rather than high-salinity media (10 to 50% artificial seawater). The greatest production of extracellular nucleic acids by P. cepacia occurred at pH 7 and 37 degrees C, suggesting that extracellular-DNA production may be a normal physiologic function of the cell. Incubation of labeled P. cepacia cells in water from Bimini Harbor, Bahamas, resulted in labeling of macromolecules of the ambient microbial population. Collectively these results indicate that (i) extracellular-DNA production by genetically altered bacteria released into aquatic environments is more strongly influenced by physiochemical factors than biotic factors, (ii) extracellular-DNA production rates are usually greater for organisms released in freshwater than marine environments, and (iii) ambient microbial populations can readily utilize materials released by these organisms.