- Volume 128, Issue 5, 1982

Three pure cultures of diatoms - a Navicula sp., a Nitzschia sp. and a Synedra sp. - grown in the presence of naphthalene at 6 or 12 °C oxidized the naphthalene to ethyl acetate-soluble and water-soluble metabolites. The major ethyl acetate-soluble metabolite was identified as 1-naphthol by gas chromatographic and mass spectral analysis. Experiments with [14C]naphthalene indicated that the extent of naphthalene metabolism ranged from 0·7 to 1·4 %.

Twenty-seven yeast species were tested for growth on methylamine and n-butylamine as sole nitrogen sources. Five species (including Saccharomyces cerevisiae) failed to grow on either amine, and a further five grew only on n-butylamine. The remainder grew on both amines. Amine oxidase activity was detected in extracts of all the strains which could grow on amines, but was generally absent from cells grown on ammonia or nitrate. From measurements in cell-free extracts of oxidase activity for a number of different amines, it is concluded that most such strains have at least two amine oxidases of different substrate specificity, and that the substrate specificity also varies between different yeast species. Based on these observations, four different groups of yeasts could be distinguished. Formaldehyde dehydrogenase activity was elevated in cells grown on amines or nitrate compared with those grown on ammonia, and activity was generally higher in cells grown on methylamine than in those grown on n-butylamine or nitrate.

Oxygen-dependent bioluminescence of Photobacterium (Vibrio) fischeri was used to measure oxygen affinities of four protozoa. The aerobic organisms Acanthamoeba castellanii and Tetrahymena pyriformis showed apparent K m values for O2 of 0·42 and 2·43 μm respectively. The aerotolerant anaerobe Tritrichomonas foetus, and the more strictly anaerobic rumen ciliate Dasytricha ruminantium, both of which have hydrogenosomes, respired with apparent K m values of 1·08 and 1·70 μm-O2. We conclude that mitochondrial respiration is not the only process conferring on organisms a high affinity for O2.

When the thiol-oxidizing agent diamide was added to a batch culture of Streptococcus faecalis during exponential growth, the specific activity of inorganic pyrophosphatase (EC 3·6·1·1) decreased to the level observed in the stationary phase. The effect of diamide was completely reversed by reduced glutathione. Furthermore, the ratio of reduced to oxidized glutathione and the specific activity of inorganic pyrophosphatase changed in a very similar fashion during batch culture. These findings, together with our earlier results, suggest that the activity of inorganic pyrophosphatase in S. faecalis is regulated by glutathione via the ratio of reduced to oxidized glutathione.

The utilization of S4O2- 6 by washed cells of Chlorobium vibrioforme f. sp. thiosulfatophilum was light-dependent. The stoichiometry for S4O2- 6 oxidation and 14CO2 assimilation was 1:3·5. Both processes were inhibited by O2, but this effect was partially reversed by flushing the reaction mixture with argon. A variety of inhibitors of thiol groups and electron transport, as well as uncouplers and ionophores, restricted these activities. During the oxidation of 35S-labelled S2O2- 3 by washed cells, the sulphone S atom (from S35SO2- 3) was rapidly oxidized to SO2- 4 via SO2- 3, whereas the first identifiable product of the oxidation of the sulphane S atom (from 35SSO2- 3) was S2- and then elemental sulphur, which was extruded into the medium and then reutilized and oxidized to SO2- 4. A number of inhibitors of thiol groups and respiratory carriers, as well as ionophores, also restricted the oxidation of S2O2- 3.

Ergosterol was identified as the major free sterol of Aspergillus nidulans by thin-layer chromatography, alumina column chromatography, gas-liquid chromatography, highperformance liquid chromatography, UV spectroscopy, proton magnetic resonance spectroscopy and mass spectral analysis. Lanosterol, the initial cyclized precursor of ergosterol, was identified as a minor component of the free sterols. In the steryl ester material, however, lanosterol was usually more abundant than ergosterol, suggesting that the esters serve as storage compounds for the membrane sterol precursors.

Transport of nutrients (glutamate, glucose and acetate) into membrane vesicles of Bacillus sp. A-007 was specifically dependent on the Na+ gradient (outside high). The nutrients were co-transported with Na+, the process being stimulated by alkaline pH. In addition to the transport process, binding of glutamate to membrane vesicles was also pH- and Na+-dependent.

N-Acetyl-β-glucosaminidase, β-glucuronidase and acid phosphatase activities were detected in cell-free extracts of Mycobacterium leprae (from armadillo liver). Extracts of bacteria which had been treated with 7-diazonaphthalene-1,3-disulphonic acid to inactivate surface enzymes retained 30–45% of the activity of the glycosidases and 15% of the activity of the acid phosphatase. When intact bacteria were treated with 1 m-NaOH, the corresponding activity in the extracts was 4–9% for the glycosidases and 7% for the acid phosphatase. Inhibition studies with lactones and the use of concanavalin A-agarose showed differences between the glycosidases in extracts of M. leprae and those of armadillo liver. Inhibition studies with vanadate using extracts from NaOH-treated bacteria and extracts of armadillo liver showed differences between the acid phosphatases. Enzymes removed from the surface of M. leprae could have been adsorbed to the surface from host tissue (i.e. lysosomal enzymes) or they could have been extracellular enzymes or associated with the bacterial membrane.

Lysine was excreted by Bacillus megaterium CII19 after growth in chemically defined media containing low concentrations of sulphate. Addition of more sulphate, or cysteine or methionine stopped lysine formation; instead ornithine was excreted. All the enzymes of the pathway leading from aspartate to lysine were assayed in extracts prepared from organisms grown in batch and continuous cultures (sulphur-or carbon-limited). The rate of the slowest step (tetrahydrodipicolinate acetylase) limited the rate of lysine synthesis to about 10 nmol min−1 (mg protein)−1, though none of the other enzymes was very much more active. Aspartokinase, aspartic β-semialdehyde dehydrogenase, dihydrodipicolinate synthase and N-acetyl-aminoketopimelate:glutamate aminotransferase were partly repressed by methionine; aspartokinase was also partly repressed by threonine. The aspartokinase repressed by methionine was inhibited by methionine plus lysine (though not by either amino acid singly), and the aspartokinase repressed by threonine was inhibited by threonine. None of the other repressible enzymes was inhibited by these amino acids. The activities of enzymes of glutamate and ornithine biosynthesis were very little altered by growth with limited or excess sulphate.

Four penicillin-binding proteins (PBPs) were observed in the soluble fraction of Caulobacter crescentus CB13, and designated PBP S1 (mol. wt 60000), PBP S2 (55000), PBP S3 (45000) and PBP S4 (40000). Caulobacter crescentus CB15, an independent isolate, possessed similar soluble PBPs, but not PBP S2. These soluble PBPs could be observed even when a cell suspension was directly reacted with [14C]penicillin G. Two of the soluble PBPs, S1 and S2, released half their bound [14C]penicillin G during a 10 min incubation, indicating that these PBPs have penicillinase-like activity. PBP S2 was very thermolabile and lost its penicillin-binding activity after incubation at 40 °C for 10 min. Mecillinam did not show selective binding to any of the Caulobacter soluble PBPs or to PBPs in the cell envelope.

Growth of the yeast Saccharomycopsis lipolytica on n-alkanes induced a hydroxylation system which, in the presence of NADPH and O2, converted lauric acid into ω-hydroxylauric acid. The specific hydroxylation activity was lower with the physiological substrate n-hexadecane than with lauric acid. This system was particulate and did not act on N-and O-dealkylated compounds. It was strongly inhibited by CO and its enzymic activity was unstable. Because of the presence of cytochrome oxidase in the preparations, spectral demonstration of the presence of cytochrome P-450 was difficult but could be achieved with the techniques developed.

By isolating phototactic mutants also impaired in thermotaxis we have confirmed that the photosensory and thermosensory transduction pathways converge in Dictyostelium discoideum slugs. Novel features of behaviour of the mutant slugs, normally obscured in ‘wild type’ strains, are the extreme bimodal distribution of directions travelled during phototaxis, and the presence of a second transition from positive to negative thermotaxis at high temperature during migration in thermal gradients. The novel behaviours result from quantitative changes in the transition temperatures during thermotaxis and the preferred angles of deviation (±α) from the direction of the light source during phototaxis. The mutant slugs orient with reduced accuracy in both phototaxis and thermotaxis. These behaviour patterns are consistent with the outcome of mutations impairing signal processing after convergence of photosensory and thermosensory transduction pathways but before adaptation processes operate to set both the preferred directions in bidirectional phototaxis and the transition temperatures in thermotaxis.

The mode of action of colicin S8 has been studied and compared with that of other colicins. Two minutes after the addition of colicin S8 to bacteria a considerable proportion of the colicin is inaccessible to trypsin. Treatment of bacteria with colicin S8 renders them more sensitive to lysis by sodium dodecyl sulphate and also inhibits motility. Like colicins K and E1, colicin S8 provokes lysis of bacteria superinfected with bacteriophage T4. Colicin S8, unlike colicin E2, prevents replication of bacteriophage T4. The incorporation of isoleucine or uracil into bacteria is inhibited by colicin S8 but, unlike colicins K and E1, the effect is multiplicity-dependent. A rapid method of titration of colicin S8 is described. The results are discussed with emphasis on the possible rearrangements at the bacterial surface and the possibility that there is more than one type of specific receptor for colicin S8.

Aspergillus nidulans has three molybdoenzvmes, nitrate reductase nitrate reductase, purine hydroxylase I and purine hydroxylase II. These three enzymes share a molybdenum-containing cofactor whose synthesis requires the inteegrity of five loci, designated cnxABC, cnxE, cnxG and cnxH. Here we report the existence of a sixth locus, designated cnxj which might be involved in the regular of cofactor levels. When grown in the presence, but not in the absence, of tungstate or methylammonium, stains carrying cnxJ1 or cnxJ2 have reduced molybdoenzyme levels as judged both from growth properties and enzyme determinations. A new cryosensitive cnxC − allele is also reported. Its phenotype at 37 °C (but not 25 °C) shows some silmiarities to that of the two cnxJ − alleles. A structural role for the cnxC (or cnxABC) product in the coafctor is tentatilvey suggested.

Streptococcus mutans chromosomal DNA cloned into the vector plasmid pBR322 in Escherichia coli is able to complement the metabolic defect of an aspartate-semialdehyde dehydrogenase (EC 1.2.1.11) gene (asd) deletion in the host strain. We constructed two Asd+ recombinant plasmids, pYA570 and pYA571, containing 4·7 and 4·5 kilobases, respectively, of S. mutans chromosomal DNA inserted into the HindIII restriction endonuclease site of pBR322 in the same orientation. The S. mutans UAB62 Asd+ DNA did not hybridize with E. coli DNA which contained an intact asd gene, but did hybridize with S. mutans UAB62 chromosomal DNA. Derivative Asd+ plasmids were then constructed from pYA570. One, pYA574, had a 4·5 kilobase S. mutans insert DNA in the opposite direction from pYA570. In another, pYA575, the S. mutans insert DNA was reduced in size to 1·3 kilobases. It was seen that the orientation of the S. mutans DNA fragment inserted into the promoter region of the pBR322 tetracycline resistance (Tcr) gene affected expression of Tcr. Orientation of the S. mutans insert also affected the stability of the plasmid in certain E. coli strains. Restriction maps for pYA570, pYA571, pYA574 and pYA575 using the endonucleases EcoRI, BamHI, HindIII, PstI and SalI were determined. Asd+ plasmid-directed protein synthesis was studied in E. coli minicells. The plasmids pYA570, pYA574 and pYA575 each produced large amounts of a protein, with a monomeric molecular weight of about 45000, that was distinct from both pBR322 and E. coli specified protein; this protein is the S. mutans asd gene product. Smaller derivatives of recombinant plasmid pYA575 that were Asd− allowed the location of the S. mutans asd gene promoter and the direction of transcription to be determined.