- Volume 79, Issue 2, 1973

SUMMARY: Mucopeptide synthesis by Pediococcus cerevisiae ATCC 8081 and P. cerevisiae crd (a methicillin-requiring substrain) was measured by incorporation of radioactivity from 14C-labelled-alanine, -glutamate, -lysine and -aspartate into the mucopeptide fraction of organisms incubated in a solution inadequate to support growth. Teichoic acid synthesis was measured by incorporation of [32P]H3PO4 into a hot trichloroacetic acid soluble fraction from isolated walls.

Mucopeptide synthesis by both strains (as measured by incorporation of [1-14C]glutamic acid) was unaffected by chloramphenicol but was sensitive to benzylpenicillin. The incorporation was resistant to methicillin in strain crd (2 mg/ml gave 50 % inhibition) but not in parent organisms (120 μg/ml gave 50 % inhibition).

Strain crd incorporated twenty times less [32P]H3PO4 into teichoic acid than did the parent strain. With either strain, benzylpenicillin (25 μg/ml) and d-cyclo-serine (200 μg/ml) inhibited this synthesis, but methicillin (500 and 1000 μg/ml) did not.

Electron micrographs of strain crd incubated for four hours with methicillin showed large deposits of wall material around septa; these were not seen in parent organisms.

SUMMARY: Synthesis and secretion of extracellular neutral and alkaline proteases in Aspergillus nidulans are repressed in the presence of low molecular weight sources of carbon, nitrogen and sulphur; protease is formed and released when the medium is deficient for any one of these elements. Induction of protease by exogenous protein does not occur.

Activation of intracellular protease is repressed by sources of carbon, nitrogen, sulphur and phosphorus, activation taking place when any one is deficient.

These two systems of repression are independent of one another and of the two previously defined systems of ammonium repression in Aspergillus nidulans. Several classes of mutant selected for ammonium derepression also have abnormal carbon repression of protease.

SUMMARY: The production of both intracellular and extracellular α-l-arabinofuranosidase (AF) by Sclerotinia fructigena was demonstrated. They had different kinetic parameters, pH stability and optimal pH values. Two peaks of activity were found on gel filtration: maximum activity was associated with the peak of lower molecular weight in the culture filtrate, but with the higher molecular weight peak in the mycelial homogenate.

Isoelectric-focusing studies on culture filtrates revealed two peaks of AF activity with pI 3.0 and 6.5 respectively, maximum activity being associated with the latter. In the mycelial homogenate, most of the activity was associated with a third peak of activity with pI 4.5.

Physical and kinetic data suggest that the two AF activities present externally in the culture filtrates are identical with the two internal ones, with corresponding pI.

SUMMARY: The effects of water potential on the photosynthesis, growth and viability of natural populations and laboratory cultures of Cyanidium caldarium were measured. C. caldarium was very sensitive to reduced water potential, matric reduction being more harmful than osmotic. The distribution of this alga in different soil areas appeared to be related to the sensitivity to water stress, larger populations being found in areas with higher water potential.

There are very steep gradients of water potential in the soils where Cyanidium caldarium is found. The water potential of the surface crust is much too low to allow growth and C. caldarium is found in a subsurface layer. The reduced light intensity below the soil surface interacts with the increased water potential to define the position of the algal layer.

SUMMARY: From an Escherichia coli strain k12 carrying a temperature-sensitive hostspecific modification (hsm) mutation, second-step mutants have been isolated that are completely deficient in modification and restriction. Complementation analysis has revealed that one group of these mutants is impaired in the specificity gene hss, while in the other group of mutant strains both mutations, i.e. the first-step temperature-sensitive and the second-step which impairs restriction and modification completely, are located in hsm. Analysis of the heterozygotes used in the complementation experiments suggested a cis- and tandem arrangement of the hs and leu genes in haploid, segregating exconjugants; however, the attachment of these genes to a cryptic plasmid was not excluded.

SUMMARY: A method of synchronous mass production of yeast zygotes is described. A yield of 30 to 40 % zygotes with the period of formation restricted to 1 h was achieved. This result was obtained by determining optimal parameters for different steps of sexual fusion of haploid a and α cells in yeast.

SUMMARY: A strain of Streptomyces coelicolor a3(2) was selected in which the wild-type autonomous SCPi plasmid had acquired an insertion of a region of the host chromosome bearing the cysB locus. This strain, called an SCPi′ strain by analogy with F′ strains of Escherichia coli, donated the cysB + allele very efficiently to a cysB recipient. The Cys+ progeny of such crosses were heterozygotes that gave rise to Cys– segregants by plasmid loss. Other donor markers, not carried on the plasmid, were donated with much lower frequencies. The SCPi-cysB plasmid was also donated rather efficiently to a strain of Streptomyces lividans, previously shown to receive the wild-type SCPi plasmid. Transfer back from S. lividans to S. coelicolor a3(2) occurred at a very much lower frequency.

SUMMARY: The histidine-induced production of a-toxin in Staphylococcus aureus strain Wood 46 declined rapidly when the pH was raised above 8.0 in the medium. This was related to the degradation of histidine by a histidase. The staphylococcal histidase was thermostable, had a Km of 1.2 × 10−2 m and pH optimum near 9.0. Both histidine and urocanic acid raised the intracellular level of histidase when added to the medium. The enzyme was inhibited by cysteine, but not by EDTA. It was slowly oxidized to an inactive form, activity being restored by glutathione or mercaptoethanol.

Histidine dipeptides had a similar or more potent stimulating effect than histidine on the production of staphylococcal a-toxin in a synthetic medium. Glycyl-l-histidine and l-alanyl-l-histidine entered the organisms more readily than free histidine. The dipeptides were hydrolysed intracellularly, and a considerable portion of the released histidine was further degraded to urocanic acid.

l-Homocarnosine (γ-aminobutyryl-l-histidine) did not stimulate α-toxin production. The uptake and intracellular hydrolysis of this dipeptide were significantly slower than those of dipeptides with the free amino group in the α-position. Accumulation of intracellular histidine could not be demonstrated without an exogenous source of histidine. This was also examined under conditions when α-toxin was produced. The results indicated that stimulation of α-toxin production was not directly correlated with the intracellular level of free histidine. Accordingly, the mediator of the stimulating effect is still unknown.

SUMMARY: There has been disagreement as to whether Neurospora crassa contains ribosomes bound to the endoplasmic reticulum. We have isolated a cellular fraction which by morphological and biochemical criteria appears to consist of membrane-bound ribosomes.

SUMMARY: Free and membrane-bound ribosomes were isolated from wild-type Neurospora crassa containing the genetic backgrund Em 5297a at 6, 12, 18, 24, 36, 48 and 72 h of growth. Log phase began at 8 h and continued to the end of 24 h, the beginning of the stationary phase. The ratio of free to membrane-bound ribosomes remained relatively constant during log phase. During stationary phase, the ratio of free to bound ribosomes rose sharply because of a decrease in the concentration of bound ribosomes.

Free ribosomes are generally assumed to synthesize intracellular proteins, whereas bound ribosomes synthesize proteins intended for secretion. These changes in membrane-bound ribosomes can be correlated with the specific activity of wallbound invertase (an intracellular enzyme) of the same strain of Neurospora. We suggest that this decrease in membrane-bound ribosomes during stationary phase is of physiological significance and occurs because of a decreasing synthesis of secreted proteins.

SUMMARY: Acid-extractable proteins were obtained from Kluyveromyces fragilis by pH titration, and partially characterized. They differ from mammalian histones and from the proteins obtained from other yeasts by similar methods. The largest fraction, extracted at pH 2.2, appeared to be predominantly of cytoplasmic origin. Quantitative changes in this fraction have been followed through the cell cycle and mapped relative to DNA synthesis, nuclear division and cell division. The total amount of such protein/cell doubled about one-third of a cycle after cell division and at a quarter of a cycle before nuclear division. DNA synthesis did not correspond to these doubling times but was almost simultaneous with cell division.

SUMMARY: Saccharomyces cerevisiae grown in glucose-limited continuous culture at low dissolved oxygen concentrations exhibited high potential respiration rates which reflected the activity of the respiratory enzyme complexes. The development of the mitochondrial respiratory enzymes, unsaturated fatty acids and lipids was highly sensitive to dissolved oxygen even in cells which were growing exclusively by fermentation. A shift to oxidative growth occurred when the oxygen concentration was increased; the apparent oxygen Km for the development of the respiratory complexes was 0.2 μ m. The increase in oxidative metabolism at increased oxygen concentration is related to the development of a functional glyoxylate cycle. Synthesis of the peroxisomal enzymes, including those of the glyoxylate cycle, occurred at significantly higher dissolved oxygen concentration than that of the mitochondrial enzymes.

SUMMARY: A complete medium of defined composition has been developed for quantitative growth of wild-type and auxotrophic mutant strains of Anacystis nidulans. This medium has proved to be more satisfactory than other complex media (for example casein hydrolysate, yeast extract) for both the isolation and the growth of auxo-trophs. Rigorous control of the pH of complete and other supplemented media is essential for quantitative growth on agar. Four diagnostic media are described which each contain a different combination of the supplements used in the complete medium and facilitate the identification of the nutritional requirements of mutants. By using these media a number of auxotrophs have been isolated including five with novel phenotypes which require respectively (i) thiamine, (ii) p-aminobenzoic acid, (iii) a combination of pyruvate or acetate plus malate or succinate or fumarate, (iv) serine or glycine and (v) adenine.

SUMMARY: Respiration of Crithidia fasciculata was more than 90 % inhibited by 0.5 mm-KCN or by a 19:1, CO:O2 gas mixture. Low-temperature difference spectra indicated the presence of a-, b- and c-type cytochromes and several CO-reacting haemoproteins. Photochemical action spectra revealed that cytochrome a 3 and another component (with an α-band in the CO-liganded state at 570 nm) act as functional terminal oxidases; no evidence was obtained for the presence of a functional cytochrome o. Extracts of the organism contain a cyanide-sensitive and CO-insensitive cytochrome c peroxidase. The possible identity of the haemoprotein(s) reacting with CO to give a Soret absorption maximum at 418 nm and previously referred to as cytochrome o is discussed.

SUMMARY: Osmotically fragile protoplasts have been prepared by the action of lytic enzymes on the mycelium of Penicillium chrysogenum and Cephalosporium acremonium. The yield of protoplasts, based on DNA content, was up to 18 %. Pretreatment of the mycelium with a thiol compound was necessary with Cephalosporium but not with Penicillium. Electron micrographs indicated that the protoplasts contained all the intracellular organelles of the mycelium and were bounded only by a cytoplasmic membrane. The metabolic activity of the protoplasts, as measured by their respiration, ability to maintain intracellular amino-acid pools, and antibiotic production, was similar to that of control mycelium. l-Valine and l-α-aminoadipic acid, which are precursors of penicillin N and cephalosporin C, were taken up and concentrated by the protoplasts of C. acremonium, although the latter transported l-valine less rapidly than did the corresponding mycelium. Protoplasts of P. chrysogenum took up l-valine but not l-α-aminoadipic acid or its δ-ester. There was little or no transport of the corresponding d-amino acids by protoplasts of either organism.

SUMMARY: A set of 121 strains of yeasts, originally described as Torulopsis, was studied by means of 63 morphological, physiological and serological characters and was subjected to numerical analysis. The matching coefficient of Sokal and Michener and the average linkage cluster analysis allowed the grouping of strains into nine phenons, formed on the 80% similarity level: stellata, dattila, colliculosa, molis-chiana, pinus–schatavii–glabrata, versatilis, gropengiesseri, wickerhamii–kruisii–candida and holmii. Resulting data agreed with published results of GC content in DNA.