Effect of l-valine and l-isoleucine on fatty acid composition of Streptomyces hygroscopicus and S. griseus

The effects of L-valine and L-isoleucine on the composition of mycelial fatty acids were investigated during growth of differentiating parent strains of Streptomyces hygroscopicus and Streptomyces griseus as well as their non-differentiating derivatives (Amy-strains) on a synthetic medium. Both in the Amy+ and Amy- strains, in the presence of L-valine, the portion of the isopalmitic acid (iC16:0) increased, but the addition of L-isoleucine led to an elevated level of the 12-methyltetradecanoic acid (aC15:0). The results suggest that the genetically determined alterations in the ratio of both fatty acids in the non-differentiating derivatives may be due to specific changes in the biosynthetic pathways of both amino acid precursors rather than due to changes of their catabolism.     

Dependence of the specific growth rate of methanogenic mutualistic cocultures on the methanogen

Methanogenesis from ethanol was studied in batch cocultures of a proton-reducing acetogenic Desulfovibrio sp. together with one of eight methanogenic bacteria representing five genera. A mathematical model of mutualistic cocultures predicts an equalisation in the specific growth rates of the two species which defines a specific growth rate for the coculture. At non-limiting ethanol concentrations the model predicts that the specific growth rate of the coculture is dependent upon the K _s (H₂) of the methanogen and its maximum specific growth rate in axenic culture when utilising H₂ as the energy source. We demonstrate experimentally that with methanogens known to have similar K _s (H₂) values, the specific growth rates of methanogenic mutualistic cocultures are dependent upon the maximum specific growth rates of the methanogens.     

Dependence of cell yield and specific growth rate ofCandida boidinii on temperature during continuous cultivation

Investigation of the dependence of cell yield ofCandida boidinii in a chemostat and specific growth rate in a turbidostat on temperature was carried out using methanol as substrate. Each of the curves obtained had only one maximum, the optimum temperature for yield (Y _s )and for specific growth rate (μ) being somewhat different. With deviation of temperature from the corresponding optimum the μ value decreases more sharply than that ofY _s . The shape of two-factor dependencesY _s (μ, T) and μ(S, T) is analyzed on the basis of these data and the known dependences ofY _s on μ and μ on the limiting substrate concentrationS. It is shown that the decelerating action of methanol on the growth rate increases at elevated and lower temperatures.     

Streptomyces hygroscopicus has two glutamine synthetase genes.

Streptomyces hygroscopicus, which produces the glutamine synthetase inhibitor phosphinothricin, possesses at least two genes (glnA and glnB) encoding distinct glutamine synthetase isoforms (GSI and GSII). The glnB gene was cloned from S. hygroscopicus DNA by complementation in an Escherichia coli glutamine auxotrophic mutant (glnA). glnB was subcloned in Streptomyces plasmids by insertion into pIJ486 (pMSG3) and pIJ702 (pMSG5). Both constructions conferred resistance to the tripeptide form of phosphinothricin (bialaphos) and were able to complement a glutamine auxotrophic marker in S. coelicolor. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of S. lividans(pMSG5) revealed a highly overexpressed 40-kilodalton protein. When GS was purified from this strain, it was indistinguishable in apparent molecular mass from the 40-kilodalton protein. The nucleic acid sequence of the cloned region contained an open reading frame which encoded a protein whose size, amino acid composition, and N-terminal sequence corresponded to those of the purified GS. glnB had a high G + C content and codon usage typical of streptomycete genes. A comparison of its predicted amino acid sequence with the protein data bases revealed that it encoded a GSII-type enzyme which had previously been found only in various eucaryotes (47 to 50% identity) and nodulating bacteria such as Bradyrhizobium spp. (42% identity). glnB had only 13 to 18% identity with eubacterial GSI enzymes. Southern blot hybridization experiments showed that sequences similar to glnB were present in all of the five other Streptomyces species tested, as well as Frankia species. These results do not support the previous suggestion that GSII-type enzymes found in members of the family Rhizobiaceae represent a unique example of interkingdom gene transfer associated with symbiosis in the nodule. Instead they imply that the presence of more than one gene encoding GS may be more common among soil microorganisms than previously appreciated.     

Influence of the growth rate on the macromolecular composition of Acinetobacter calcoaceticus in carbon-limited chemostat culture

Abstract Infectious phage particles can be formed in vitro when extracts of T1-infected cells are incubated with T1 DNA. The DNA packaging system is based on mixtures of complementing extracts from Escherichia coli sup⁰ cells infected with the amber mutants am 4 (gene 16) or am 10 (gene 13). Gene 16 mutants are defective in the formation of DNA-filled heads but make proheads; gene 13 mutants are defective in prohead formation. Three forms of DNA have been packaged: (1) endogenous concatemeric DNA present in mixtures of am 4 and am 10 mutant extracts; (2) concatemeric DNA; (3) virion DNA both when supplied exogenously to mixtures of am 4 · am 20 and am 10 · am 20 double mutant extracts ( am 20 inhibits T1 DNA synthesis). The reaction requires added ATP, Mg²⁺ and spermidine for optimum efficiency and produces about 1.5 × 10³ pfu/ μ g and about 1 × 10⁴ pfu/ μ g for exogenous concatemeric and virion DNA, respectively.     

The induction of antibiotic synthesis in Saccharopolyspora erythraea and Streptomyces hygroscopicus by growth rate decrease is accompanied by a down-regulation of protein synthesis rate

Abstract The relationship between antibiotic production and culture growth rate in Saccharopolyspora erythraea and Streptomyces hygroscopicus was manipulated by changing the growth-limiting substrate. Carbon- and nitrogen-limited cultures were studied and antibiotic synthesis was obtained in both cases in Saccharopolyspora erythraea cultures and in nitrogen-limited Streptomyces hygroscopicus cultures. In all cultures where antibiotic was detected, onset of antibiotic production coincided with the minimal protein synthesis rate. Further investigation in Saccharopolyspora erythraea cultures indicated that this corresponded to minimum ratio of charged to uncharged tRNA, i.e. when uncharged tRNA accumulated. This latter phenomenon was investigated in the presence of a protein synthesis inhibitor.     

Morphological engineering of Streptomyces hygroscopicus var. geldanus: regulation of pellet morphology through manipulation of broth viscosity

Actinomycetes, especially members of the genus Streptomyces, are responsible for producing the majority of known antibiotics. The production of antibiotics by filamentous organisms is often dependent on the morphology and size distribution of the pellet population within the culture. Particle interaction and subsequent pellet formation are primarily dependent on the rate of collision of particles in culture, which is in turn, a function of fluid turbulence. The microbial polysaccharide xanthan gum was used to artificially regulate the apparent viscosity (_a) of S. hygroscopicus fermentation broths with the aim of controlling particle interaction, aggregation and hence pellet formation. An increase in both pellet count and biomass concentration from approximately 2,000 to 8,000 pellets ml^–1 and 0.9–2.1 g l^–1 dry weight of biomass, as well a decrease in the mean pellet volume from 0.014 to 0.004 mm³ was observed in cultures supplemented with 3 g l^–1 xanthan gum. The addition of xanthan gum significantly alters fluid rheology by increasing the _a. Counter-intuitively, an increase in the _a within the experimental range examined resulted in an increase in the rate of gas–liquid mass transfer. This was attributed to the predominantly diffusive nature of oxygen transfer in shake flask cultures.     

Dependence of the composition of the avermectin complex of Streptomyces avermitilis on glucose content of the growth medium

The effects of glucose concentration in the medium and O-methyl-L-threonine resistance on the ratio of components of the avermectin complex produced by Streptomyces avermitilis have been studied. Glucose deficiency increases the ratio of components A and a, while decreasing that of components 1 in the complex. A mutation that renders the microorganisms resistant to O-methyl-L-threonine (an analog of isoleucine) increases the ratio of components a, while decreasing that of components 1 in the complex. The distribution of a and b in fractions 1 and 2 remains constant: the values of the ratio a/b in the fractions amount, respectively, to 1:1 and 2:1. The relation of the variations in the composition of the avermectin complex to changes in the carbohydrate metabolism of the producer stain, underlain by availability of the source of carbon, is discussed.     

Formation and regeneration of protoplasts of Streptomyces hygroscopicus

Techniques are described for the production and regeneration of protoplasts of Streptomyces hygroscopicus and related strains. No single technique was successful in producing protoplasts from all strains. Regeneration of protoplasts to the mycelial growth form was greatly improved by modifying the physical regeneration environment. Protoplast formation and regeneration was achieved in six out of the seven strains studied.     

Dependence of the specific growth rate of Escherichia coli MI 30 on the ammonia concentration]

The transient behaviour of ammonium limited continuous cultures of E. coli ML 30 led to the hypothesis that the bistability of pyruvate formation primarily is caused by a bistability of the ammonia metabolism. Therefore, a function of mu([NH+4]) should be expected different from that of Monod type. Measurements of the specific growth rate during washout of continuous cultures at different ammonium concentrations and at such low cell concentrations that the changes in the ammonium concentration of the medium could be neglected, showed a complex function with a relative minimum near 2 mg/1NH+4. This function allows bistability of the ammonium concentration in an ammonium limited continuous culture. The results are discussed on the basis of the two systems of ammonia assimilation found in prokaryotic cells.     

Pentalenolactone I and hygromycin A, immunosuppressants produced by Streptomyces filipinensis and Streptomyces hygroscopicus.

Immunosuppressants isolated from Streptomyces filipinensis and S. hygroscopicus were identified with pentalenolactone I and hygromycin A, respectively. The compounds as well as cyclosporin A showed immunosuppressant activity in the mixed lymphocyte reaction, and pentalenolactone I and cyclosporin A suppressed IL-2 production, however, hygromycin A did not. Hygromycin A may have immunosuppressant activity by a different mechanism from pentalenolactone I, cyclosporin A and tacrolimus.     

Changes in macromolecular composition and morphology of Bacteroides fragilis cultured in a complex medium.

Changes in cell macromolecular composition during different phases of growth correlated with alterations in morphology of the obligately anaerobic bacterium, Bacteroides fragilis, grown in a complex medium.     

Distribution functions of variables characterizing the mycelial morphology of Streptomyces hygroscopicus grown in glucose-limited chemostat cultures

The distribution of variables characterizing the morphology of the mycelium of Streptomyces hygroscopicus grown in glucose-limited chemostat cultures at different specific growth rates were investigated statistically. The values of the hyphal growth unit (L/N) and the values of the distance from the apex to the first branch (Lp) are normally distributed, but the values of the distance between neighbouring branches are logarithmically normal distributed. The distribution functions are discussed from the biological point of view.     

Establishment of a system suitable for analysis of balanced and unbalanced growth of Streptomyces hygroscopicus

A small-scale system was developed in which balanced growth of Streptomyces hygroscopicus occurred. Although the balanced growth, verified by corresponding increase of ATP, DNA, RNA, protein and mycelial length, was restricted to a relatively short period of the life cycle, it lasted for at least two doublings. The conditions for balanced growth could be altered by various treatments to induce imbalance. This system could be applied to study regulation in the mycelium under well-defined conditions.