bluuebunny/crossref_metadata_2025 huggingface dataset:
Glucose transport in isolated prosthecae of Asticcacaulis biprosthecum
Authors: Larson R J , Pate J L
Published: January 2020

Active transport of glucose in prosthecae isolated from cells of Asticcacaulis biprosthecum was stimulated by the non-physiological electron donor N, N, N', N'-tetramethyl-p-phenylenediamine dihydrochloride. Glucose uptake was mediated by two transport systems; the apparent Km of the high-affinity system was 1.8 muM and that of the low-affinity system was 34 muM. Free glucose accumulated within prosthecae at a concentration 60 to 200 times above that present externally, depending on the Km of the system being observed. The glucose transport system in prosthecae was stereospecific for D-glucose, and neither methyl alpha-D-glucopyranoside nor 2-deoxyglucose was transported. Uptake of glucose was inhibited by N-ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB), and the inhibition by PCMB but not by NEM was reversed by dithiothreitol. Glucose uptake was also inhibited by the uncoupling agents 5-chloro-3-t-butyl-2'-nitrosalicylanilide (S-13), 5-chloro-3-(p-chlorophenyl)-4'-chlorosalicylanilide (S-6), and carbonyl-cyanide m-chlorophenylhydrazone (CCCP) and by the respiratory inhibitor KCN. Efflux of glucose from preloaded prosthecae was induced by PCMB and KCN, but not by S-13 or CCCP. Glucose uptake was not affected by arsenate or an inhibitor of membrane-bound adenosine triphosphatases, N, N'-dicyclohexylcarbodiimide. The lack of inhibition by these two compounds, combined with the extremely low levels of adenosine 5'-triphosphate present in prosthecae, indicates that adenosine 5'-triphosphate is not involved in the transport of glucose by prosthecae.
Undermethylated transfer ribonucleic acid from a relaxed strain of Bacillus subtilis: construction of the strain and analysis of the transfer ribonucleic acid
Authors: Keisel N , Vold B
Published: January 2020

A strain of Bacillus subtilis is described from which undermethylated transfer ribonucleic acid (tRNA) can be obtained. The tRNA's from a methionine-limited culture were compared with those from a control culture with respect to general nucleoside composition, methylated components, and amino acid acceptor activity. The undermethylated tRNA's had the normal amounts of the four major nucleosides, pseudouridine, and 5-methyluridine (ribothymidine), but were deficient in methylated nucleosides other than 5-methyluridine. These methyl-deficient nucleosides can be fully remethylated in the presence of the appropriate methylases. Since the majority of the work characterizing undermethylated tRNA's has been done using Escherichia coli, the work with B. subtilis presents some interesting comparisons and offers an alternative substrate for methylase studies.
Molecular nature of two beta-lactamase-specifying plasmids isolated from Haemophilus influenzae type b
Authors: De Graaff J , Elwell L P , Falkow S
Published: January 2020

The molecular nature of two beta-lactamase-specifying plasmids isolated from two separate ampicillin-resistant Haemophilus influenzae type b strains was examined. A 30 X 10(6)-dalton (30-Mdal) plasmid (RSF007) had a copy number of approximately 3 per chromosomal equivalent and a mole fraction guanine plus cytosine content of 0.39. By heteroduplex analysis the 30-Mdal plasmid was found to contain the entire ampicillin translocation DNA segment (TnA) found on R factors of enteric origin. A 3.0-Mdal plasmid (RSF0885) was found as a multicopy pool of approximately 28 copies per chromosomal equivalent, had a mole fraction guanine plus cytosine content of 0.40, and contained only about one-third of the transposable TnA sequence. RSF007 and RSF0885 appeared to be unrelated plasmids in that they share base sequence homology only within the confines of the TnA segment. The 3.0-Mdal Haemophilus plasmid was used to transform E. coli to ampicillin resistance but was found to be unstable in this host in the absence of antibiotic. The possibility that R-plasmids arose in Haemophilus by the translocation of TnA from a donor R-factor onto an indigenous H. influenzae plasmid is discussed.
Mode of replication of the conjugative R-plasmid RSF1040 in Escherichia coli
Authors: Crosa J H , Luttropp L K , Falkow S
Published: January 2020

Replicating deoxyribonucleic acid (DNA) molecules of plasmid RSF1040, a deletion mutant of the conjugative R plasmid R6K, appear in the electron microscope as partially supercoiled structures with two open circular branches of equal size, although open structures with three branches, two branching points and no supercoiled regions (theta structures) were also found at a lower frequency. The partially supercoiled molecules sediment more rapidly than native covalently closed circular DNA in neutral sucrose gradients and band at a position intermediate between covalently closed circular and open circular DNA in CsClethidium bromide gradients. Electron microscope measurements of the linear EcoRI-treated replicative intermediates indicate that replication can be initiated at two sites (origins) on the plasmid DNA molecule located at about 23% (alpha) and 39% (beta) of the total genome length from an EcoRI end designated arbitrarily as the "left-hand" end of the molecule. The overall replication of RSF1040 is asymmetrically bidirectional. Replication from the alpha origin proceeds first to the "right" to a unique termination site located some 55% of the total genome length from the left-hand end of the molecule. At this point replication proceeds from the alpha origin to the "left" (i.e., opposite to the original direction of replication) until replication of the molecule is completed. Replication also proceeds from the beta origin asymmetrically to the unique terminus site.
Formation of the formate-nitrate electron transport pathway from inactive components in Escherichia coli
Authors: Scott R H , DeMoss J A
Published: January 2020

When Escherichia coli was grown on medium containing 10 mM tungstate the formation of active formate dehydrogenase, nitrate reductase, and the complete formate-nitrate electron transport pathway was inhibited. Incubation of the tungstate-grown cells with 1 mM molybdate in the presence of chloramphenicol led to the rapid activation of both formate dehydrogenase and nitrate reductase, and, after a considerable lag, the complete electron transport pathway. Protein bands which corresponded to formate dehydrogenase and nitrate reductase were identified on polyacrylamide gels containing Triton X-100 after the activities were released from the membrane fraction and partially purified Cytochrome b1 was associated with the protein band corresponding to formate dehydrogenase but was not found elsewhere on the gels. When a similar fraction was prepared from cells grown on 10 mM tungstate, an inactive band corresponding to formate dehydrogenase was not observed on polyacrylamide gels; rather, a new faster migrating band was present. Cytochrome b1 was not associated with this band nor was it found anywhere else on the gels. This new band disappeared when the tungstate-grown cells were incubated with molybdate in the presence of chloramphenicol. The formate dehydrogenase activity which was formed, as well as a corresponding protein band, appeared at the original position on the gels. Cytochrome b1 was again associated with this band. The protein band which corresponded to nitrate reductase also was severely depressed in the tungstate-grown cells and a new faster migrating band appeared on the polyacrylamide gels. Upon activation of the nitrate reductase by incubation of the cells with molybdate, the new band diminished and protein reappeared at the original position. Most of the nitrate reductase activity which was formed appeared at the original position of nitrate reductase on gels although some was present at the position of the inactive band formed by tungstate-grown cells. Apparently, inactive forms of both formate dehydrogenase and nitrate reductase accumulate during growth on tungstate which are electrophoretically distinct from the active enzymes. Activation by molybdate results in molecular changes which include the reassociation of cytochrome b1 with formate dehydrogenase and restoration of both enzymes to their original electrophoretic mobilities.
Threonine deaminase from Escherichia coli: feedback-hypersensitive enzyme from a genetic regulatory mutant
Authors: Calhoun D H
Published: January 2020

A mutation, ilvA538, in the gene coding for the biosynthetic L-threonine deaminase of Escherichia coli K-12 has previously been demonstrated to have pleiotropic regulatory effects leading to low and invariant expression of some of the isoleucine-valine biosynthetic enzyme, and altered expression of the branched-chain aminoacyl-tRNA synthetases. Strain PS187, which carries the ilvA538 allele, has a partial growth requirement for L-isoleucine and is characterized by a sensitivity to growth inhibition by L-leucine. The experiments reported here demonstrate that the L-threonine deaminase produced by strain PS187 is hypersensitive to inhibition by the pathway end product L-isoleucine. In addition, L-leucine, which acts at relatively high concentrations in vitro as an inhibitor of L-threonine deaminase from the wild type, is a more potent inhibitor of the activity of the mutant enzyme. Forty-six derivatives of strain PS187 were isolated as spontaneous mutants resistant to the growth-inhibitory effects of L-leucine. Two of these, strains MSR14 and MSR16, produce an L-threonine deaminase that is more resistant than the wild type to L-isoleucine inhibition, and intermediate between the wild type and strain PS187 with respect to L-leucine inhibition. Strains MSR14 and MSR16 produce L-threonine deaminase and dihydroxyacid dehydrase, the ilvD gene product, at the low levels characteristic of the parent strain. Other L-leucine-resistant derivatives of strain PS187 produce higher levels of the feedback-hypersensitive L-threonine deaminase. Thus, the sensitivity to growth inhibition by L-leucine observed with strain PS187 appears to be related both to the hypersensitivity of L-threonine deaminase to inhibition of catalytic activity and to the low level of ilv gene expression. The results reported here indicated that L-threonine deaminase is structurally altered in strain PS187, and thus provide further support for the proposal that L-threonine deaminase participates as a genetic regulatory element for the expression of the branched-chain amino acid biosynthetic enzymes.
Comparative studies on membrane-associated, folded chromosomes from Escherichia coli
Authors: Dworsky P
Published: January 2020

Nuclear bodies were isolated from Escherichia coli spheroplasts by two different lysis procedures. Their lipid and protein content and the superhelix density was measured. The preparations differed mainly in respect to the amount of the attached membrane material, which seems to be an essential factor in maintaining the stability of the nuclear bodies. The superhelix density was found to be higher than that of naturally occurring, covalently closed, circular deoxyribonucleic acids. The influence of the preparative method on the superhelicity was comparatively small.
Recognition sites for chemotactic repellents of Bacillus subtilis
Authors: Ordal G W
Published: January 2020

Repellents of Bacillus subtilis include many membrane-active compounds, such as uncouplers of oxidative phosphorylation, local anesthetics, chlorpromazine (a central nervous system depressant), and tetraphenylboron (a lipophilic anion). Normally, bacteria swim smoothly, and occasionally tumble, but addition of repellent causes all bacteria to tumble, then later resume original frequency of swimming and tumbling (adaptation). Bacteria adapted to repellent can then be tested to determine the minimum concentration (threshold) of the same or different repellents that causes tumbling. The results indicate that repellents act at (saturable) recognition sites, which differ for chemically different species. An implication is that uncouplers of oxidative phosphorylation affect cell properties by interaction at specific locations.
Hypersensitivity to Catabolite Repression in the
l
-Arabinose Operon of
Escherichia coli
B/r Is
trans
Acting
Authors: Sheppard David E. , Eleuterio Marianne
Published: January 2020

Mutations causing hypersensitivity to catabolite repression have been assigned to gene
araC
(activator protein) by complementation analysis. The
araO
(operator region) is non-essential for catabolite repression.
Inhibition of dimethyl ether and methane oxidation in Methylococcus capsulatus and Methylosinus trichosporium
Authors: Patel R , Hou C T , Felix A
Published: January 2020

Metal-chelating or -binding agents inhibited the oxidation of dimethyl ether and methane, but not methanol, by cell suspensions of Methylococcus capsulatus and Methylosinus trichosporium. Evidence suggests that the involvement of metal-containing enzymatic systems in the initial step of oxidation of dimethyl ether and methane.
Extragenic suppression of two ribosomal protein cistrons lying near the rif locus in Escherichia coli
Authors: Molholt B
Published: January 2020

The experiments reported here involve temperature-sensitive mutations in two of five cistrons encoding 50S ribosomal proteins that lie near the rif locus in Escherichia coli. I selected spontaneous TS+ mutants able to grow at elevated temperatures in which the TS+ event takes place outside this tract of cistrons near rif. Six distinct classes of extragenic suppressors were found, five of which have been mapped. Two of these suppressors lie near 64 min, a region known to be rich in cistrons ribosomal proteins (Dennis and Nomura, 1975). The remaining three extragenic suppressors were located near 16.5, 47, and 86 min.
Ribonucleic acid synthesized in meiotic cells of Saccharomyces cerevisiae: effect of culture medium pH
Authors: Curiale M S , Petryna M M , Mills D
Published: January 2020

Pulse-labeled ribonucleic acid (RNA) was extracted from polysomes of sporulating cells of Saccharomyces cerevisiae and characterized in sucrose gradients and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Transfer RNA, ribosomal RNA, and heterodisperse RNA, presumed to be messenger RNA, were synthesized during a 20-min pulse at T4 and T6 when labeling was performed in sporulation medium adjusted to pH 6.0. Furthermore, ribosomal RNA was processed into functional ribosomes during the pulse. The specific activity of pulse-labeled RNA of cells labeled in sporulation medium where the pH was unadjusted at T4 (pH 7.8) and T9 (pH 8.6) was 20- to 50-fold lower than RNA from cells labeled at pH 6.0. The low specific activity resulted from a 50-fold reduction in uptake of labeled precursors when the medium pH was greater than 7.2. However, heterodisperse RNA ranging from 4-17S in size and transfer RNA were synthesized during the pulse at T4 (pH 7.8),but the low specific activity of ribosomal RNA prevented a thorough analysis of its synthesis. Cellular impermeability at T9 (pH 8.6) resulted in minimal uptake of label, and an analysis of pulse-labeled transcripts was impossible. A comparison of the percantage of polysomal material indicate, however, that these cells were at least as active in translation as cells pulse-labeled at pH 6.0.
Tryptophan operon regulation in interspecific hybrids of enteric bacteria
Authors: Manson M D , Yanofsky C
Published: January 2020

We examined tryptophan regulation in merodiploid hybrids in which a plasmid carrying the trp operon of Escherichia was introduced into Trp mutants of other enteric genera, or in which a plasmid carrying the trpR+ (repressor) gene of E. coli was transfered into fully constitutive trpR mutants of other genera. In these hybrids the trp operon of one species is controlled by the repressor of a different species. Similar investigations were possible in transduction hybrids in which either the trp operon or the trpR+ locus of Shigella dysenteriae was introduced into E. coli. Our measurements of trp enzymes levels in repressed and nonrepressed cells indicate that Trp regulation is normal, with only minor quantitative variations, in hybrids between E coli and Shigella dysenteriae, Salmonella typhimurium, Klebsiella aerogenes, Serratia marcescens, and Proteus mirabilis. Our results support the idea that a repressor-operator mechanism for regulating trp messenger ribonucleic acid production evolved in a common ancestor of the enteric bacteria, and that this repressor-operator recognition has been conversed during the evolutionary divergence of the Enterobacteriaceae.
Lack of glucose phosphotransferase function in phosphofructokinase mutants of Escherichia coli
Authors: Roehl R A , Vinopal R T
Published: January 2020

Phosphofructokinase (pfkA) mutants of Escherichia coli are impaired in growth on all carbon sources entering glycolysis at or above the level of fructose 6-phosphate (nonpermissive carbon sources), but growth is particularly slow on sugars, such as glucose, which are normally transported and phosphorylated by the phosphoenolpyruvate, (PEP)-dependent phosphotransferase system (PTS).
Ultrastructure of the cell walls of two closely related clostridia that possess different regular arrays of surface subunits
Authors: Sleytr U B , Glauert A M
Published: January 2020

Cell walls of Clostridium thermohydrosulfuricum and C. thermosaccharolyticum have a two-layered structure, consisting of a thin, lysozyme-sensitive murein layer and a surface (S) layer composed of hexagonally or tetragonally arranged subunits. The subunits can be removed from the murein layer by treatment of cell wall preparations, are composed of a fragile, pH-sensitive monolayer of macromolecular subunits. In both organisms the first stage of the cell division process involves only the plasma membrane and the murein layer. During the subsequent cell separation, a surplus of S-layer subunits appears at the site of division, and consequently the newly formed cell poles remain completely covered by the s layer throughout the separation process. In autolyzed cells an additional layer of subunits assembles on extended areas of the inside of the mucopeptide layer. These observations indicate that the biological function of the S layer depends on its ability to maintain a complete covering of the cell surface at all stages of cell growth and division.
Phosphatase of Chlamydomonas reinhardi: biochemical and cytochemical approach with specific mutants
Authors: Matagne R F , Loppes R , Deltour R
Published: January 2020

The unicellular alga Chlamydomonas reinhardi produces two constitutive acid phosphatases and three depressible phosphatases (a neutral and two alkaline ones) that can utilize napthyl phosphate as a substrate. Specific mutants depressible phosphatase were used to investigate biochemical properties and the cytochemical localization of these enzymes. The two constitutive phosphatases show similar pH optima (about 5.0) and Km values (2 x 10(-3) to 3.3 x 10(-3) M) but differ in their heat sensitivity and affinity for glycerophosphate.
Cellular organization of Bacillus subtilis: sodium dodecyl sulfate-induced cell partitioning into zebra structures
Authors: Mendelson N H , Haag S M , Cole R M
Published: January 2020

Cells of Bacillus subtilis heated in high concentrations of sodium dodecyl sulfate (5%) and then washed free of detergent with a hot salt solution (80 C) become structurally reorganized into regions of densely compacted cytoplasm (termed zebras) and regions of sparsely filled material (termed spaces). Size distribution studies of zebras indicate that division-suppressed mutants and wild-type cells both yield zebras of comparable length. Similarly the lengths of zebras found in populations emerging from spores are uniform in one-, two-, three-, and four-zebra-containing cells. In contrast, the length of spaces is slightly larger than that of zebras and is unusually large in two-zebra-containing cells. The locations of zebras and spaces along cell length have been studied in spore out-growth populations. A statistical procedure developed previously in genome location investigations was used to analyze the location of zebras along cell length. The data indicate that as cells elongate, new sites arise where the cell contents are strongly bound to the cell surface. Within filament populations produced by division-suppressed mutants there is a linear relationship of mean filament length and zebra number per filament. These data indicate that cytoplasm in filaments with no obvious structural compartmentalizations may be organized into units associated with particular regions of cell surface. The attachment of cell contents to the cell surface may involve deoxyribonucleic acid. Zebra-containing cells digested with proteolytic enzyme and ribonuclease are converted to cells that contain a crystalline-like granule fixed at the location of each zebra. Exposure to deoxyribonuclease mobilizes these granules within the cell wall.
Transport of glucose and glycine in Schizosaccharomyces pombe during the cell cycle
Authors: Kubitschek H E , Claymen R V
Published: January 2020

Cell growth and uptake of glucose and glycine during the cell cycle were studied in synchronous cultures of Schizosaccharomyces pombe. Rates of accumulation of glucose and glycine were constant during most of the cell cycle, implying a constant rate of cell mass increase. Rates of uptake of glycine appeared to double at an average cell age of 0.9 generations.
Effects of crp mutations on adenosine 3',5'-monophosphate metabolism in Salmonella typhimurium
Authors: Rephaeli A W , Saier M H
Published: January 2020

Wild-type Salmonella typhimurium could not grow with exogenous cyclic adenosine 3',5'-monophosphate (AMP) as the sole source of phosphate, but mutants capable of cyclic AMP utilization could be isolated provided the parental strain contained a functional cyclic AMP phosphodiesterase.All cyclic AMP-utilizing mutants had the growth and fermentation properties of cyclic AMP receptor protein (crp) mutants, and some lacked cyclic AMP binding activity in vitro. The genetic defect in each such mutant was due to a single point mutation, which was co-transducible with cysG. crp mutants isolated by alternative procedures also exhibited the capacity to utilize cyclic AMP. crp mutants synthesized cyclic AMP at increased rates and contained enhanced cellular cyclic AMP levels relative to the parental strains, regardless of whether or not cyclic AMP phosphodiesterase was active. Moreover, adenylate cyclase activity in vivo was less sensitive to regulation by glucose, possibly because the enzyme II complexes of the phosphotransferase system, responsible for glucose transport and phosphorylation, could not be induced to maximal levels. This possibility was strengthened by the observation that enzyme II activity (measured both in vitro by sugar phosphorylation and in vivo by sugar transport and chemotaxis) was inducible in the parental strain but not in crp mutants. The results suggest that the cyclic AMP receptor protein regulates cyclic AMP metabolism as well as catabolic enzyme synthesis.
Influence of pH on the rate of ribosomal ribonucleic acid synthesis during sporulation in Saccharomyces cerevisiae
Authors: Wejksnora P , Haber J E
Published: January 2020

The rate of synthesis of ribosomal RNA (rRNA) is much slower during sporulation than during vegetative growth of yeast. If sporulating cells are transferred from normal incubation conditions at pH 8.8 to the same medium adjusted to pH 7.0, the rate of rRNA synthesis increased to approach that observed in vegetative cells. The response to the pH change is quite rapid, occurring within 10 min. THE PH-dependent, rate-limiting step appears to be in the processing of 35S ribosomal precursor RNA to the final 26S and 18S RNA species. A similar pH effect also was found for the rate of protein synthesis. However, no change in respiration was observed when the pH was lowered. These results indicate that the observed differences in rate of rRNA synthesis in vegetative and sporulating cells are a consequence of pH and are not intrinsic to sporulation. The results also support the correlation between rRNA processing and protein synthesis.
Isolation and identification of the conidial germination factor of Neurospora crassa
Authors: Horowitz N H , Charlang G , Horn G , Williams N P
Published: January 2020

The germination-essential substance (germination factor [GF]) that is lost from conidia of Neurospora crassa on exposure to solutions of low water activity has been isolated and identified as a group of iron-transport compounds, or siderochromes. The principal siderochrome of conidia is ferricrocin, a cyclic hexapeptide. A closely related substance, ferrichrome C, is tentatively identified as a minor constituent. The same substances are also present in extracts of mycelium along with small amounts of a third siderochrome, which has not been identified. The GF activity of culture filtrates is due to coprogen, the only siderochrome previously identified with N. crassa.
Adenosine 5'-triphosphate synthesis energized by an artificially imposed membrane potential in membrane vesicles of Escherichia coli
Authors: Tsuchiya T , Rosen B P
Published: January 2020

Adenosine 5'-triphosphate (ATP) synthesis driven by an artificially imposed membrane potential in right-side-out membrane vesicles of Escherichia coli was investigated. Membrane vesicles prepared in the presence of adenosine diphosphate were loaded with K+ by incubation with 0.5 M potassium phosphate. Addition of valinomycin resulted in the synthesis of 0.2 to 0.3 nmol of ATP/mg of membrane protein, whereas no synthesis was observed after addition of nigericin. Addition of K+, dicyclohexylcarbodiimide, carbonylcyanide p-trifluoromethoxyphenylhydrazone, or azide to the assay buffer inhibited ATP synthesis. Adenosine diphosphate and Mg2+ were found to be required. Ca2+, which can replace Mg2+ for the hydrolytic activity of the Mg2+-adenosine triphosphatase (ATPase) (EC 3.6.1.3), could not replace Mg2+ in the synthetic reaction and, in fact, inhibited ATP synthesis even in the presence of Mg2+. Strain NR-70, a mutant lacking the Mg2+-ATPase, was unable to synthesize ATP using an artificially imposed membrane potential. Additionally, the Mg2+-ATPase was found to contain tightly bound ATP.
Natural paucity of anaplerotic enzymes: basis for dependence of Arthrobacter pyridinolis on L-malate for growth
Authors: Krulwich T A , Sharon B I , Perrin L S
Published: January 2020

Previous work has shown that in Arthrobacter pyridinolis the transport systems for glucose and several amino acids are respiration coupled, with malate oxidation occurring concomitantly with transport. The requisite malate has to be supplied exogenously, so that growth on glucose or certain amino acids only occurs if malate is also present in the medium. These and other data suggested that A. pyridinolis might be deficient in anaplerotic enzymes, which maintain intracellular levels of dicarboxylic acids. A comparative study was undertaken of anaplerotic enzymes in A. pyridinolis and in a closely related species, A. crystallopoietes, which has respiration-coupled transport of glucose but can grow on glucose without added malate. The paucity of anaplerotic enzymes in A. pyridinolis and its probable relationship to the malate requirement for growth on glucose were documented as follows: (i) A. crystallopoietes, but not A. pyridinolis, possesses phosphoenolpyruvate carboxylase activity, and neither species contains pyruvate carboxylase; (ii) both A. pyridinolis and A. crystallopoietes possess glyoxylate pathways that are induced by acetate but not by hexoses; (iii) isocitrate lyase-deficient mutants of A. pyridinolis fail to grow on rhamnose and fructose as well as acetate; and (iv) mutants of A. crystallopoietes that require malate for growth on glucose are deficient in phosphoenolpyruvate carboxylase.
R factor-mediated polarized chromosomal transfer in Escherichia coli C
Authors: Hedén L O , Rutberg L
Published: January 2020

Five transferable drug resistance factors (R factors) with the ability to bring about chromosomal transfer in Escherichia coli C were investigated with respect to the direction and origin of chromosome transfer. They were found to constitute two groups, both with a clockwise direction of transfer. One group has its origin of transfer between arg and pro, and the other group has its origin of transfer between try and man on the E. coli C chromosome. With one R factor in particular, the rapid increase in the production of recombinants was followed by a decrease in numbers as mating was prolonged.
Ultrastructural characterization of capsulated Haemophilus influenzae type b and two spontaneous nontypable mutants
Authors: Doern G V , Buckmire F L
Published: January 2020

Capsulated Haemophilus influenzae type b and two spontaneous mutants (classes I and II variants) were characterized by transmission and scanning electron microscopy. When cells were treated with type b-specific antiserum prior to manipulations for electron microscopy, sectioned capsulated cells had electron-dense, fibrous capsular antigen-antibody complexes around them. In negatively stained preparations, the complexes appeared as electron-transparent zones surrounding cells. In contrast, only residual electron-dense, extracellular material was seen in sectioned, untreated, capsulated cells, and electron-dense "bridges" connected adjacent cells in negatively stained preparations. No extracellular capsular material was seen around the class I and II variants. Characteristic electron-translucent regions were always observed within the cytosol of the class I cells, both in thin sections and by negative staining. These areas were located adjacent to the cell envelope separating the plasma membrane from the dense cytoplasmic matrix. At times, electron-dense, thread-like material extended from the dense cytoplasmic matrix to the plasma membrane. No such regions were seen in the capsulated and class II cells. Class I cells fixed with methanol or suspended in NaCl or phosphate-buffered saline prior to treatment with fluorescein-tagged type b-specific antiserum (FTA reagent) exhibited, by immunofluorescence, patches of capsular antigen along their sides. However, when fixed with glutaraldehyde or OsO4 or suspended in tris-(hydroxymethyl)aminomethane plus Ca2+ buffer prior to treatment with FTA reagent, no patches of capsular antigen were seen. Subsequent exposure of the latter cells to methanol followed by treatment with FTA reagent resulted in the reappearance of the patches of capsular antigen. Thus, in the class I variant the capsular antigen is unlikely to be surface located. Scanning electron microscopy revealed that class I and II variant cells within undisturbed colonies were regularly aligned side-by-side, whereas cells within colonies of the capsulated strain were randomly distributed.
Repair of deoxyribonucleic acid in ultraviolet light-sensitive and -resistant Dictyostelium discoideum strains
Authors: Guialis A , Deering R A
Published: January 2020

Some responses of the cellular slime mold Dictyostelium discoideum to ultraviolet light (UV) irradiation were investigated by analyzing two aspects of deoxyribonucleic acid (DNA) excision repair in the vegetative cells: (i) the fate of thymine-containing dimers and (ii) the production and rejoining of single-strand breaks. Experiments were done with the parental, radiation-resistant NC-4 strain and with the radiation-sensitive gammas-13 strain. The majority (greater than 85%) of the thymine-containing dimers produced in both strains by an energy fluence of 100/Jm2 were removed from the acid-insoluble DNA fraction within the first 3 to 4 h of reincubation in the dark. Moreover, as measured by alkaline sucrose gradients, single-strand breaks appeared in the DNA of both NC-4 and gammas-13 irradiated cells very rapidly and at low temperatures. This was presumed to be a result of the incision (nicking) step of excision repair as performed by UV-specific endonuclease(s). In NC-4 the time required for dimer excision correlated with the sealing of breaks as well as with the UV-induced division delays. In gammas-13 the single-strand breaks were closed at a slower rate than in NC-4. However, this was not accompanied by more extensive division delays.
Morphogenic effects of alpha-factor on Saccharomyces cerevisiae a cells
Authors: Lipke P N , Taylor A , Ballou C E
Published: January 2020

Saccharomyces cerevisiae mating type a cells enlarged and elongated when exposed to alpha-factor, a sex pheromone produced by mating-type alpha cells. This morphogensis required exogenous-D-glucose, nitrogen, and phosphate, and cells in exponential phase responded better than stationary-phase cells. Morphogenesis was blocked by cycloheximide and by inhibitors of cell wall biosynthesis such as 2-deoxy-D-glucose, 2-deoxy-2-fluoro-D-glucose, and 2-deoxy-2-fluoro-D-mannose, but not by polyoxin D. One to two hours after addition of pheromone, a cells became more susceptible to lysis by glucanases, a change that was dependendent on the dose of alpha-factor and was blocked by drugs that block morphogenesis. On the other hand, treatment with alpha-factor did not increase susceptibility to attack by trypsin, subtilisin, or exo-alpha-mannanase. Radioactive label, incorporated into cell wall polysaccharides during treatment with alpha-factor, was not secreted into the medium during morphogenesis. Analysis of the labeled wall polymers showed that alpha-factor-treated cells contain more glucan and less mannan than control cells, and that the mannan of treated cells contains an increased proportion of shorter side chains and unsubstituted backbone mannose units. Thin-section electron microscopy of treated cells revealed that the cell wall possesses a diffuse outer layer in the extension and is thinner at the tip.
Fragility of plasma membranes in Saccharomyces cerevisiae enriched with different sterols
Authors: Hossack J A , Rose A H
Published: January 2020

Saccharomyces cerevisiae NCYC 366, grown under strictly anaerobic conditions to induce requirements for an unsaturated fatty acid (supplied by Tween 80) and a sterol, contained free sterol fractions enriched to the extent of 67 to 93% with the exogenously supplied sterol (campesterol, cholesterol, 7-dehydrocholesterol, 22, 23-dihydrobrassicasterol, beta-sitosterol, or stigmasterol). Cells enriched in any one of the sterols did not differ in volume, growth rate, contents of free sterol, esters and phospholipids, or phospholipid composition. Cholesterol-enriched cells contained about 2% more lipid than cells enriched in any of the other sterols, which was largely accounted for by increased contents of triacylglycerols and, to a lesser extent, esterified sterols. Phospholipids were enriched to the extent of about 52 to 63% with C18:1 residues. Cells enriched in ergosterol or stigmasterol were slightly less susceptible to the action of a wall-digesting basidiomycete glucanase than cells enriched with any one of the other sterols. The capacity of the plasma membrane to resist stretching, as indicated by the stability and volume of spheroplasts suspended in hypotonic solutions of buffered sorbitol (particularly in the range 0.9 to 0.7 M), was greater with spheroplasts enriched in sterols with an unsaturated side chain at C17 (ergosterol or stigmasterol) than with any of the other sterols. Plasma membranes were obtained from spheroplasts enriched in cholesterol or stigmasterol and had free sterol fractions containing 70 and 71%, respectively, of the sterol supplied exogenously to the cells. The sterol-phospholipid molar ratios in these membranes were, respectively, 1:7 and 1:8.
Effects of 4-[beta-(diethylamino)-ethoxy]-benzophenone upon carotenogenesis in Rhodospirillum rubrum
Authors: Hayman E P , Yokoyama H
Published: January 2020

Carotenoid production was determined in illuminated anaerobically maintained cultures of Rhodospirillum rubrum in media with and without 4-[beta-(diethylamino)-ethoxy]-benzophenone. In treated cultures, lycopene--which normally is not produced by R. rubrum--accumulated as the predominant pigment, and total carotenoids increased five- to sixfold.
Low-frequency, pbsi-mediated plasmid transduction in Bacillus pumilus
Authors: Bramucci M G , Lovett P S
Published: January 2020

Three bacteriophages were tested for ability to transduce the plasmid of pPL10 between W mutant derivatives of Bacillus pumilus NRS 576. Phage PBP1- and PMB1-generated plasmid transductants occurred at about 10% the frequency of transductants for a chromosome marker. Phage PBS1-generated plasmid transductants occurred at less than 0.1% the frequency of transductants for a chromosome marker. Possible reasons for the extremely reduced capacity of PBS1 to generate plasmid transductants are discussed.
Properties of pGC1, a lac plasmid orginating in Yersinia enterocolitica 842
Authors: Cornelis G , Bennett P M , Grinsted J
Published: January 2020

pGC1, a self-transmissible lac plasmid that originated in Yersinia enterocolitica 842, is described. This plasmid is freely transmissible between strains of Y. enterocolitica and from such strains to strains of Escherichia coli. The plasmid can also be transferref among G. coli strains and back to Y. enterocolitica, although at a greatly reduced frequency. pGC1 has a molecular weight of about 33.0 x 10(6), has a base composition of about 44 mol% guanine plus cytosine, and is fi-.
Outer membrane of Escherichia coli K-12: isolation of mutants with altered protein 3A by using host range mutants of bacteriophage K3
Authors: Manning P A , Puspurs A , Reeves P
Published: January 2020

A series of mutants has been isolated with alterations to protein 3A of the outer membrane. These mutations map at the previously described con locus as shown by cotransduction with pyrD. Most of them do not have detectable levels of protein 3A but are thought to have low levels of altered protein. These mutants have been detected by screening con mutants, isolated as resistant to bacteriophage K3, for their ability to plaque host range mutants of this bacteriophage. These host range phage mutants have activity spectra on the various con mutants that enable the bacterial mutants to be arranged in an order of increasing resistance to the host range phage mutants, from mutants sensitive to all host range phage to those sensitive to only one class. Likewise, the phage can be arragned in an order of increasing ability to plaque on the con mutants. Some of the mutants resemble the previously described con mutants in being tolerant to colicins K and L, and others resemble them in being highly defective as recipients with the F factor. These properties vary independently, suggesting that protein 3A can be modified to independently affect the three properties of bacteriophage receptor function, involvement in colicin sensitivity, and involvement in conjugation.
Metabolism of 5-hydroxylysine in Pseudomonas fluorescens
Authors: Friede J D , Henderson L M
Published: January 2020

Hydroxylysine is metabolized via two routes by a Pseudomonas fluorescens strain as shown by the oxidation of selected intermediates. Hydroxy-L-lysine is oxidized via a pathway analogous to the monooxygenase pathway for L-lysine, and data suggest that at least some of tthe enzymes are those involved in the metabolism of L-lysine. Hydroxy-L-lysine is also converted by a racemase to allohydroxy-D-lysine, which is then degraded via a pathway analogous to, but different from, that described for D-lysine, involving hydroxy-L-pipecolate, 2-amino-5-hydroxyadipate, and 2-hydroxyglutarate. Data obtained with mutants unable to oxidize L-pipecolate suggest that the enzymes for the metabolism of hydroxy-L-pipecolate are distinct from those for L-pipecolate. Studies on D- and L-lysine degradation have shown that the previously described pathways for these compounds are present in this soil pseudomonad.
Temperature-sensitive ribonucleic acid polymerase mutant of Salmonella typhimurium with a defect in the beta' subunit
Authors: Young B S , Guterman S K , Wright A
Published: January 2020

Localized mutagenes of Salmonella typhimurium followed by a [3H]uridine enrichment procedure yielded a temperature-sensitive strain with a mutation in the rpo region of the chromosome. Ribonucleic acid (RNA) polymerase (EC 2.7.7.6; nucleoside triphosphate: RNA nucleotidyltransferase) purified from this mutant was considerably less active at the nonpermissive temperature than wild-type enzyme. Furthermore, the enzyme from this mutant, unlike RNA polymerase of previously isolated temperature-sensitive mutants, was as thermostable as wild-type enzyme when preincubated at 50 degrees C. Subunit reconstitution experiments have shown that the temperature sensitivity is caused by an alteration in the beta' subunit of the enzyme.
Ferric hydroxamate transport without subsequent iron utilization in Bacillus megaterium
Authors: Arceneaux J E , Byers B R
Published: January 2020

Iron transport and utilization were examined in Bacillus megaterium Ard1, a mutant that is resistant to the hydroxymate antibiotic A22765 and whose growth is inhibited by the structurally similar hydroxamate Desferal. Rapid, low-level uptake of Desferal-50Fe was observed; such uptake was temperature and energy independent. Gel filtration chromatography of the cytoplasmic fraction of protoplasts labeled with Desferal-55Fe for 30 to 120 s demonstrated only unchanged esferal-55Fe in the cytoplasm. Although B. megaterium Ard1 showed transport of Desferal-59Fe by a process that resembles facilitated diffusion, this organism was unable to transfer iron from this chelate to cellular macromolecules for metabolic use. High-level transport of the ferric hydroxamate schizokinen-59Fe by B. megaterium Ard1 was both temperature and energy dependent. Within 30 s, protoplasts labeled with schizokinen-55Fe contained iron associated with certain macromolecules and in an apparent "pool" of schizokinen-55Fe in the cytoplasmic fraction. Prior transport of Dseferal-55Fe by protoplasts of strain Ard1 did not interfere with subsequent transport and utilization of schizokinen-59Fe. These studies suggest that transport of ferric hydroxamates may occur by a facilitated diffusion-type process; transfer of iron to cellular macromolecules may drive high-level transport of the chelate and may be the step at which energy is required in the iron transport-assimilation process.
Three-dimensional reconstruction of whole cells of Streptococcus faecalis from thin sections of cells
Authors: Higgins M L
Published: January 2020

A new ultrastructural technique has been developed to study the geometry of cell wall assembly in Streptococcus faecalis, which is believed to occur between pairs of raised bands located on the organism's surface. Three-dimensional reconstructions of these new regions of envelope growth are produced from the mathematical rotation (around a central axis) of various measurements taken from central, longitudinal thin sections of cells. These reconstructions can be used to calculate the surface area and volume of the septal and peripheral walls that were supposedly present in any given cell before sectioning. In an accompanying paper, it is shown how such surface and volume estimations, coupled with other measurements of length, thickness, and curvature, can be used to characterize a cycle of envelope growth in this organism. The validity of the assumptions used to reconstruct cells by rotation and the possible sources of error in using this technique are discussed.
Study of cycle of cell wall assembly in Streptococcus faecalis by three-dimensional reconstructions of thin sections of cells
Authors: Higgins M L , Shockman G D
Published: January 2020

A new ultrastructural method was used to study rounds of envelope synthesis that occur in Streptococcus faecalis in "growth zones" found between pairs of naturally occurring surface markers. The technique consists of producing three-dimensional reconstructions of these growth zones from the mathematical rotation, about a central axis, of measurements taken from central, longitudinal thin sections of cells. A cycle of exponential-phase envelope growth was then simulated by arranging a series of these reconstructions in increasing order of the amount of peripheral wall surface area or the amount of cell volume that each was calculated to contain. Using this simulated cycle of growth, the geometry of a single growth zone during a round of synthesis was studied. Based on this analysis, a model was developed for the assembly of the cell wall of S. faecalis. The model states that new cell wall surface is synthesized by the regulated flow of essentially two channels of cell wall precursors into a single growth zone. One channel of precursors would be involved in the assembly of a bilayered cross wall that would proceed at a fairly constant rate until the cross wall closes. The second channel of precursors would be involved in the separation of the bilayered cross wall into two segments of peripheral wall. These precursors would intercalate into and thicken the separating layers of the cross wall. The flow of precursors through this channel would be progressively reduced through a cycle. These decreases, when coupled with internal hydrostatic pressure, apparently would result in the enlarging peripheral wall becoming increasingly more curved and would also promote cell division by reducing the total amount of cell wall that must be assembled in order for septation to occur.
Organization of the nucleoplasm in Escherichia coli visualized by phase-contrast light microscopy, freeze fracturing, and thin sectioning
Authors: Woldringh C L , Nanninga N
Published: January 2020

The organization of the nucleoplasm in Escherichia coli was studied by comparing the results obtained by freeze fracturing and thin sectioning. In addition to exponentially growing cells, we used chloramphenicol-treated cells which show a well-defined nucleoplasm, in the phase-contrast light microscope and can therefore function as a control for treatments necessary for electron microscopy. Two factors were found to determine the visibility of the nucleoplasm in freeze fractures: first, the state of lateral aggregation of deoxyribonucleic and fibrils, which is enhanced by postfixation with OsO4 according to the Ryter-Kellenberger technique; second, the presence of ice crystals. When their formation is prevented by the use of high concentration of freeze-protecting agents, the nucleoplasm appears as a smooth region in cells that have been prefixed. In unfixed cells, however, the freeze-protecting agent causes disappearance of the nucleoplasm by rearrangement of structures within the cell. This observation makes it hard to determine whether the deoxyribonucleic acid in vivo dispersed, as found after glutaraldehyde prefixation, or compact, as after OsO4 prefixation.
Ultrastructure and elemental composition of dormant and germinating Diplodia maydis spores
Authors: Murphy J A , Thompson M R , Pappelis A J
Published: January 2020

The ultrastructure of Diplodia maydis spores was studied in thin sections with a transmission electron microscope. Storage vacuoles were evenly distributed in the two cells. Some of the vacuoles that contained a dense osmiophilic sphere(s) were surrounded by a membrane, and had membranous aggregates around their periphery. The sport wall was composed of an electron-dense layer and an electron-translucent layer. An inner cytoplasmic membrane was present. Dormant and germinating spores were studied with scanning electron microscopy and also with a Si (Li) energy-dispersive X-ray analyzer. The dormant spore was ovate and usually two-celled with a central septum. Germination proceeded via a germ tube from the side of one end of the cell. Of several methods for preparation of specimens for X-ray analysis studied, freeze-dried spores mounted on carbon stubs and then further carbon coated gave the best results. X-ray analyses revealed that spore populations contained large amounts of Si, P, Cl, and K, smaller amounts of S and Ca, and trace amounts of Mg and Al. Analyses of single spores revealed high K and Cl and low P and Mg at one end of the cell with concomitant low K and Cl and high P and Mg in the central portion and other end of the cell. In two-celled germinating spores, high K and Cl occurred in the end of the nongerminating spore cell, whereas the germinating cell contained high P and Mg and low K and Cl. X-ray image maps revealed that K and Cl were located together at one end of the spore.