bluuebunny/crossref_metadata_2025 huggingface dataset:
Studies with a Gluconate-6-Phosphate Dehydrogenase Mutant in
Escherichia coli
Strain C
Authors: Robbins R. S. , Kelly Beatrice
Published: January 2020

A map location of the gluconate-6-phosphate dehydrogenase (
gnd
) marker was estimated in
Escherichia coli
C at approximately 46 min by P1 transduction. The
gnd
locus appears to lie between the co-transducible histidine and prophage P2 location I markers.
Correlation of Growth Inhibition Patterns to Nucleoside Transport Models in
Neurospora crassa
Authors: Magill Jane M. , Magill Clint W.
Published: January 2020

Growth of inhibition patterns provide evidence for a common nucleoside transport or utilization system, a separate system or systems for adenine transport, and another adaptable mechanism of adenosine transport.
In Vitro Formation of Nitrate Reductase Using Extracts of the Nitrate Reductase Mutant of
Neurospora crassa, nit-1
, and
Rhodospirillum rubrum
Authors: Ketchum Paul A. , Sevilla Cynthia L.
Published: January 2020

In vitro formation of reduced nicotinamide adenine dinucleotide phosphate (NADPH)–nitrate reductase (NADPH: nitrate oxido-reductase, EC 1.6.6.2) has been attained by using extracts of the nitrate reductase mutant of
Neurospora crassa, nit-1
, and extracts of either photosynthetically or heterotrophically grown
Rhodospirillum rubrum
, which contribute the constitutive component. The in vitro formation of NADPH-nitrate reductase is characterized by the conversion of the flavin adenine dinucleotide (FAD) stimulated NADPH-cytochrome
c
reductase, contributed by the
N. crassa nit-1
extract from a slower sedimenting form (4.5
S
) to a faster sedimenting form (7.8
S
). The 7.8
S
NADPH-cytochrome
c
reductase peak coincides in sucrose density gradient profiles with the NADPH–nitrate reductase, FADH
2
–nitrate reductase and reduced methyl viologen (MVH)–nitrate reductase activities which are also formed in vitro. The constitutive component from
R. rubrum
is soluble (both in heterotrophically and photosynthetically grown cells), is stimulated by the addition of 10
−4
M Na
2
MoO
4
and 10
−2
M NaNO
3
to cell-free preparations, and has variable activity over the pH range from 3.0 to 9.5. The activity of the constitutive component in some extracts showed a threefold stimulation when the pH was lowered from 6.5 to 4.0. The constitutive activity appears to be associated with a large molecular weight component which sediments as a single peak in sucrose density gradients. However, the constitutive component from
R. rubrum
is dialyzable and is insensitive to trypsin and protease. These results demonstrate that
R. rubrum
contains the constitutive component and suggests that it is a low molecular weight, trypsin- and protease-insensitive factor which participates in the in vitro formation of NADPH nitrate reductase.
Basic Amino Acid Transport in
Escherichia coli
: Properties of Canavanine-Resistant Mutants
Authors: Rosen Barry P.
Published: January 2020

A mutant of
Escherichia coli
strain CanR 22 has been isolated which is resistant to growth inhibition by canavanine, an analogue of arginine. The properties of this strain and of another canavanine-resistant mutant, JC182-5 (isolated by Celis et al. [5]), were studied. The mutation is pleiotropic in that it results in a reduction in the activity of two distinct permeases, the arginine-specific and lysine-arginine-ornithine transport systems. The lesion maps at min 56 of the
E. coli
linkage map, at or near the
arg
P locus. Although strain CanR 22 excretes arginine, this excretion appears to result from reduced ability to concentrate arginine, rather than the loss of transport ability being the result of excretion. This conclusion is based on findings with a canavanine-resistant strain auxotrophic for arginine, which exhibits transport properties similar to those of the prototrophic strains. Additionally, growth in the presence of arginine or ornithine results in a repression of the activity of the two basic amino acid transport systems. Neither the arginine-specific nor the lysine-arginine-ornithine binding proteins of the mutant cells show significant alterations in terms of amount, physical properties, or kinetic parameters. These observations lead to the proposal of a model for the two basic amino acid transport systems in which two carrier proteins with different specificities interact with a common energy coupling mechanism. A lesion in the gene (or one of the genes) for this coupling mechanism can confer canavanine resistance.
Stable Ribonucleic Acid Synthesis in Stringent (
rel
+
) and Relaxed (
rel
−
) Polyamine Auxotrophs of
Escherichia coli
K-12
Authors: Srinivasan P. R. , Young Delano V. , Maas Werner
Published: January 2020

The relationship of polyamines to stable ribonucleic acid (RNA) synthesis under conditions of amino acid withdrawal or chloramphenicol treatment was examined with the use of a closely related
rel
+
,
rel
−
pair conditionally incapable of synthesizing putrescine. Under conditions of polyamine starvation, the cellular sperimidine level fell to one-third to one-half of the value observed in putrescine-supplemented cultures and putrescine became undetectable; cadaverine was synthesized by both strains, but the relaxed strain, MA 252, accumulated less cadaverine per cell than its stringent twin, MA 254. Upon amino acid withdrawal, the stringent strain remained stringent whether starved of or supplemented with polyamines. Similarly, the relaxed strain was capable of making RNA either with or without polyamine starvation. On the addition of chloramphenicol or upon amino acid withdrawal in the relaxed strain, supplementation with spermidine had no effect on the initial rate of RNA synthesis, although RNA accumulation was greater in the presence of added spermidine. Spermidine added at the conclusion of RNA synthesis prompted additional synthesis, although preincubation with spermidine again had no effect on the initial rate. All forms of stable RNA species were made with polyamine supplementation. The present data appear to rule out the possibility that polyamines are primary causative agents in stimulating RNA synthesis, but rather suggest an indirect or secondary role for spermidine in which the polyamines “stimulate” stable RNA synthesis probably by relieving RNA product inhibition of RNA synthesis.
Development of Ultraviolet Resistance in Sporulating
Bacillus cereus
T
Authors: Germaine G. R. , Coggiola Elizabeth , Murrell W. G.
Published: January 2020

Ultraviolet (UV) resistance patterns of sporulating
Bacillus cereus
were determined. Six different categories of UV resistance were discernible as the organism progressed from exponential growth to the free, mature spore. Light microscope observations allowed the assignment of certain sporulation stages to specific UV resistance categories. Marked changes in UV resistance were associated with stages III, mid-IV, and post-IV of sporulation. Dipicolinic acid was shown to sensitize forespores to UV radiation. Mechanisms invoked to explain the different UV categories are discussed.
Occurrence of Cell Lytic Enzymes in Blue-Green Bacteria
Authors: Ingram L. O.
Published: January 2020

Detergent extracts of three blue-green bacteria (
Agmenellum quadruplicatum
strain BG1,
Anacystis nidulans
strain TX20, and
Nostoc
sp. strain MAC) contained enzymes capable of lysing suspensions of
Micrococcus lysodeikticus
. The enzyme preparation from
A. quadruplicatum
released soluble reducing fragments from purified peptidoglycan. The lytic activity exhibited a pH optimum between 6 and 7, was relatively heat stable, and was susceptible to attack by proteolytic enzymes. These results extend the range of bacterial types exhibiting cell lytic activity as well as confirm the existence of the lytic system commonly observed in “water blooms”.
Characterization of
Escherichia coli
Mutants Tolerant to Bacteriocin JF246: Two New Classes of Tolerant Mutants
Authors: Foulds John , Barrett Constance
Published: January 2020

Several hundred independent bacteriocin-tolerant mutants have been isolated without mutagenesis from three strains of
Escherichia coli
. On the basis of patterns of sensitivity to eight different colicins, over 85% of these mutants could be grouped into four classes. Two classes of mutants, class A and class B, are equivalent to
tolA
and
tolB
type mutants. We found
tolA
and
tolB
mutants were sensitive to the antibiotic bacitracin. The other two classes of bacteriocin-tolerant mutants, class F and class G, are distinguished from other types of colicin-tolerant mutants on the basis of sensitivity to colicins, dyes, detergents, antibiotics, and chelating agents. The mutation in class F and class G mutants is located between 21 to 23 min on the
E. coli
chromosome. We propose to designate the loci of these mutations as
tolF
and
tolG
, respectively.
Production and Characterization of the Slime Polysaccharide of
Pseudomonas aeruginosa
Authors: Evans Leigh R. , Linker Alfred
Published: January 2020

The slime polysaccharides produced by
Pseudomonas aeruginosa
isolated from a variety of human infections were investigated. Slime production in culture seemed optimal when adequate amounts of carbohydrate were present and under conditions of either high osmotic pressure or inadequate protein supply. The polysaccharides produced by the organisms were similar to each other, to the slime of
Azotobacter vinelandii
, and to seaweed alginic acids. They were composed of β-1,4-linked
d
-mannuronic acid residues and variable amounts of its 5-epimer
l
-guluronic acid. All bacterial polymers contained
o
-acetyl groups which are absent in the alginates. The polysaccharides differed considerably in the ratio of mannuronic to guluronic acid content and in the number of
o
-acetyl groups. The particular composition of the slime was not found to be characteristic for the disease process from which the mucoid variants of
P. aeruginosa
were obtained.
Isolation of Hfr Strains from R
+
and ColV2
+
Strains of
Escherichia coli
and Derivation of an R′
lac
Factor by Transduction
Authors: Nishimura Akiko , Nishimura Yukinobu , Caro Lucien
Published: January 2020

Temperature-resistant revertants were isolated from
Escherichia coli
strains carrying a temperature-sensitive
dnaA
mutation (initiation of chromosome replication) and either a repressed or a derepressed F-like R factor or a ColV2 factor. Many of the revertants had all the properties of Hfr strains, with a variety of directions and origins of transfer. From one such revertant, episomes carrying the R factor and part of the
lac
region (R′
lac
) could be isolated by transduction. This system offers a good selection for Hfr strains produced by integration of various episomes and for the isolation of R′ factors.
Purification and Regulation of Glucose-6-Phosphate Dehydrogenase from
Bacillus licheniformis
Authors: Opheim Dennis , Bernlohr R. W.
Published: January 2020

d
-Glucose-6-phosphate nicotinamide adenine dinucleotide phosphate (NADP) oxidoreductase (EC 1.1.1.49) from
Bacillus licheniformis
has been purified approximately 600-fold. The enzyme appears to be constitutive and exhibits activity with either oxidized NAD (NAD
+
) or oxidized NADP (NADP
+
) as electron acceptor. The enzyme has a pH optimum of 9.0 and has an absolute requirement for cations, either monovalent or divalent. The enzyme exhibits a

K
m

of approximately 5 μM for NADP
+
, 3 mM for NAD
+
, and 0.2 mM for glucose-6-phosphate. Reduced NADP (NADPH) is a competitive inhibitor with respect to NADP
+
(

K
m

= 10 μM). Phosphoenolpyruvate (

K
m

= 1.6 mM), adenosine 5′-triphosphate (

K
m

= 0.5 mM), adenosine diphosphate (

K
m

= 1.5 mM), and adenosine 5′-monophosphate (

K
m

= 3.0 mM) are competitive inhibitors with respect to NAD
+
. The molecular weight as estimated from sucrose density centrifugation and molecular sieve chromatography is 1.1 × 10
5
. Sodium dodecyl sulfate gel electrophoresis indicates that the enzyme is composed of two similar subunits of approximately 6 × 10
4
molecular weight. The intracellular levels of glucose-6-phosphate, NAD
+
, and NADP
+
were measured and found to be approximately 1 mM, 0.9 mM, and 0.2 mM, respectively, during logarithmic growth. From a consideration of the substrate pool sizes and types of inhibitors, we conclude that this single constitutive enzyme may function in two roles in the cell—NADH production for energetics and NADPH production for reductive biosynthesis.
Conjugal Deoxyribonucleic Acid Replication by
Escherichia coli
K-12: Effect of Chloramphenicol and Rifampin
Authors: Fenwick R. G. , Curtiss Roy
Published: January 2020

Conjugal replication of R64-11 deoxyribonucleic acid (DNA) and the concomitant transfer of R64-11 DNA to DNA-deficient minicells are dependent upon processes that are inhibited by rifampin and chloramphenicol. The rifampin-sensitive product is not present in vegetatively growing cells and is needed to initiate both conjugal DNA replication in donor cells and DNA transfer to recipient minicells. If the rifampin-sensitive product is a ribonucleic acid (RNA) molecule (rather than RNA polymerase itself), our data indicate that this RNA species required for initiation of conjugal activity does not need to be translated into a protein product. The chloramphenicol-sensitive product(s) is present in vegetatively growing cells in sufficient quantity to permit most donor cells to carry out one round of plasmid conjugal replication and transfer. The initiation of second and subsequent rounds of conjugal replication and transfer are dependent on the synthesis of both the rifampin-sensitive and chloramphenicol-sensitive products. Our results demonstrate a correspondence between the amount of conjugal DNA replication in the donor and the amount of DNA transferred to recipient minicells under all conditions, and therefore suggest but do not prove that plasmid transfer is dependent on conjugal DNA replication. The results also add additional proof that R64-11 transfer to minicells is discontinuous. All of these results are discussed in regard to further refinements of old models for the mechanism of conjugal transfer as well as a more radical departure from current dogma.
Production of Volatile Nitrogenous Compounds from the Degradation of Streptomycin by
Pseudomonas maltophilia
Authors: Fenton John J. , Harsch Harold H. , Klein Dolph
Published: January 2020

Ammonia, methylamine, and pyridine were detected in broth filtrates of a streptomycin-degrading strain of
Pseudomonas maltophilia
during growth on streptomycin as a sole carbon and nitrogen source. Ammonia and methylamine, quantitatively measured by conversion to chromophores with picryl sulfonic acid, were found to accumulate in broth, whereas pyridine concentration increased in the early stages of streptomycin degradation and then decreased as the degradation of the antibiotic neared completion. Exogenous pyridine was metabolized by washed-cell suspensions. Use of
N
-streptomycin-
methyl
-
14
C
showed that the methylamine arose from the
N
-
l
-glucosamine-methyl moiety of streptomycin. Methylamine was an end product and was not further metabolized by cells.
Thermal Death of a Hydrocarbon Bacterium in a Nonaqueous Fluid
Authors: Severance M. M. , LaRock P. A.
Published: January 2020

A hydrocarbon-utilizing
Brevibacterium
which grew into the oil phase of an oil-water system was tested for survival at elevated temperature. Cells suspended in oil and cells that had been resuspended in aqueous solution were tested by placing 1-ml samples of the cell suspension in small test tubes immersed in a controlled-temperature water bath. The resultant survival curves in oil consisted of two parts, a flat shoulder obtained in the first half of the heating period, followed by a break indicating rapid die-off. The break in the curves occurred after 50% of the cells were killed. This occurred at exposures of 25, 15, and 8 min for 78, 88.6, and 96.2 C, respectively. The survival curve for 63.5 C in the aqueous solution was a rapid, exponential die-off. The actual increase in survival of the organism in oil is reflected by the length of the shoulder portion. The shoulder occurs only in an oil medium and is increased by decreasing temperature and increasing age of the culture.
Death Resulting from Fatty Acid Starvation in Yeast
Authors: Henry Susan A.
Published: January 2020

Mutants of
Saccharomyces cerevisiae
having the genotypes
fas
1 (fatty acid synthetase minus) and
fas
1,
ole
1 (fatty acid synthetase and fatty acid desaturase minus) were found to undergo logarithmic death when deprived of required fatty acids, whereas
ole
1 strains did not. During the first 2 to 3 h of fatty acid starvation, macromolecular synthesis occurred at apparently normal rates, although cell division stopped by the end of the 1st h. Cell death commenced at approximately the 2nd to the 3rd h, and within 24 h, depending upon conditions, 2 to 4 log orders of death had occurred. The loss of viability was accelerated by the addition of detergent, but could be largely prevented by the interruption of protein synthesis, either by amino acid starvation or by the use of cycloheximide. The possible significance of this phenomenon in terms of membrane biosynthesis is discussed.
Fine Structures of Cell Envelopes of
Chlamydia
Organisms as Revealed by Freeze-Etching and Negative Staining Techniques
Authors: Matsumoto Akira
Published: January 2020

The cell walls of
Chlamydia psittaci
(meningopneumonitis strain) were examined by the freeze-etching and negative staining techniques. It was observed that the cleaved convex surface of the developmental, reticulate body was covered with numerous non-etchable particles 9 to 10 nm in diameter, these particles being rarely seen on the concave surface. Similarly, the convex surface of the mature, elementary body (EB) was covered with many particles but the concavity lacked these particles. After etching, the smooth concave surface of the EB appeared to have a hexagonally arrayed subunit structure, on which the button structure (B structure) was observed. Each B structure had a diameter of 27 nm and several B structures were grouped together in a hexagonal arrangement with a center-to-center spacing of 45 nm. In a limited area of the negatively stained EB cell wall, hexagonally arrayed rosette structures were present, with a center-to-center spacing similar to the B structures seen in the freeze-etched preparation. Each rosette, about 19 to 20 nm in diameter, appeared to be composed of a radial arrangement of nine subunits. The freeze-fractured cell wall-cytoplasmic membrane complexes indicated that the outer surface of the cytoplasmic membrane which appeared as the convex surface was covered with the fine particles, and thus it was likely that frozen EB was cleaved at the gap between the cell wall and ctyoplasmic membrane. On the cleaved inclusion, several groups of fine particles were observed. In each group, the particles were arranged hexagonally with the spacing ranging from 20 to 50 nm.
Membranes of
Rhodopseudomonas spheroides
: Interactions of Chromatophores with the Cell Envelope
Authors: Niederman Robert A.
Published: January 2020

Under carefully controlled ionic conditions, large-scale preparations of highly purified chromatophores and cell envelopes were obtained from phototrophically grown
Rhodopseudomonas spheroides
by zonal ultracentrifugation. The majority of the bacteriochlorophyll a was located in a single, discrete chromatophore band, whereas the envelopes were nearly devoid of photopigment. The envelope fraction contained substantial quantities of succinic dehydrogenase and cytochromes, confirming that phototrophically grown cells contain a photopigment-deficient cytoplasmic membrane. Magnesium at concentrations of 1.0 mM or higher caused chromatophores to reversibly aggregate with the cell envelope. Significant aggregation was also promoted by other divalent metals (Co
2+
> Mn
2+
> Ca
2+
> Mg
2+
), but aggregation was less extensive with monovalent cations. These results account for the distribution of photopigments in two bands reported by others and further suggest that the photosynthetic apparatus of
R. spheroides
is located on membranes largely distinct from the cell wall-cytoplasmic membrane complex.
Surface-Bound Nuclease of
Staphylococcus aureus
: Purification and Properties of the Enzyme
Authors: Okabayashi Kinji , Mizuno Den'ichi
Published: January 2020

The surface-bound nuclease of
Staphylococcus aureus
liberated during formation of protoplasts was purified 1,000-fold by chromatography on phosphocellulose. Its properties were compared with those of the known extracellular nuclease, purified 200-fold by the same procedures. The adsorbance of the surface-bound nuclease on phosphocellulose was distinctly different from that of the extracellular nuclease, but other properties of the two enzymes were similar. Both enzymes had a pH optimum of about 10 and required Ca
2+
for activity. Both enzymes hydrolyzed deoxyribonucleic acid (DNA) and ribonucleic acid, and denatured DNA was a better substrate than native DNA. Both enzymes were inhibited by the same metal ions. Nuclease-less mutants of
S. aureus
were isolated from
S. aureus
209P by using
N
-methyl-
N
′-nitroso-
N
-nitrosoguanidine. These mutants contained neither surface-bound nor extracellular nuclease activity. These results suggest that the surface-bound and extracellular nucleases are expressed from the same cistron of
S. aureus
.
Non-Continuous Translation of Coat Protein and Ribonucleic Acid Polymerase Cistrons in MS2 Bacteriophage Ribonucleic Acid
Authors: Grubman Marvin J. , Nakada Dai
Published: January 2020

In an MS2 phage ribonucleic acid (RNA)-directed in vitro protein-synthesizing system, the coat protein cistron and the adjacent RNA polymerase cistron are translated non-continuously. The ribosomes which have completed the synthesis of coat protein dissociate from the MS2 RNA and do not read through the intercistronic gap. Translation of the adjacent RNA polymerase cistron requires ribosomes other than those translating the coat protein cistron.
Studies of Respiratory Components and Oxidative Phosphorylation in Mitochondria of mi-1
Neurospora crassa
Authors: Drabikowska Alicja , Kosmakos Frank C. , Brodie Arnold F.
Published: January 2020

Oxidative phosphorylation has been demonstrated with mitochondria of the mi-1 respiratory mutant of
Neurospora crassa
. The P/O ratios observed with these mitochondria were approximately 0.8 with citrate and 0.4 with either externally added reduced nicotinamide adenine dinucleotide (NADH), succinate, or ascorbate-tetramethyl-
p
-phenylenediamine (TPD). These P/O ratios suggest that there are only two sites of phosphorylation in mitochondria isolated from young (20 to 24 h) cultures of the mi-1 mutant. The energy-dependent reduction of NAD
+
with succinate and the phosphorylation associated with ascorbate-TPD oxidation indicate that the first and the third sites of energy coupling are present in this mutant. Difference spectra of mitochondria from young cultures of the mi-1 mutant revealed the presence of cytochrome
c
. Cytochromes
b
and
a
+
a
3
were not detected. However, in the presence of antimycin A, a small peak in the Soret region at 430 nm was observed. A carbon monoxide difference spectrum revealed the presence of a component of the respiratory chain with a spectrum similar to that of cytochrome
o
. It is of interest that respiratory inhibitors such as antimycin A, 2-
n
-nonylhydroxyquinoline
N
-oxide, and cyanide abolished phosphorylation but only partially inhibited oxidation. It is postulated that the mi-1 respiratory system contains two pathways of electron transport—the first is associated with a phosphorylating pathway, whereas the second is a non-phosphorylating electron transport pathway.
Mechanism of Carbamyl Phosphate Inhibition of Nitrogenase of
Clostridium pasteurianum
Authors: Seto Belinda L. , Mortenson L. E.
Published: January 2020

Carbamyl phosphate caused a maximal inhibition of 50% of the in vitro nitrogenase activity measured by acetylene reduction and dinitrogen reduction. The addition of 1 mM carbamyl phosphate to a N
2
-fixing culture caused a rapid decrease of 30% of the acetylene reduction activity and also repression of nitrogenase biosynthesis. However, carbamyl phosphate had no effect on the reductant-dependent adenosine triphosphate hydrolysis and H
2
evolution reactions catalyzed by nitrogenase. Studies on the binding of carbamyl phosphate to nitrogenase and each of its two components (azoferredoxin and molybdoferredoxin) indicated that optimal binding was obtained only in the presence of an operating nitrogenase system. Moreover, the binding seemed to be on the molybdoferredoxin component rather than azoferredoxin. From a Scatchard plot and a reciprocal plot of the data, the values of
n
= 2 and dissociation constant (
K
) of approximately 5 × 10
−5
M were obtained. The value for the dissociation constant was of the same order of magnitude as the endogenous level of carbamyl phosphate in a N
2
-fixing cell. The carbamyl phosphate pool in NH
3
-grown cells was twice that of N
2
-fixing cells.
Regulation of a Ligand-Mediated Association-Dissociation System of Anthranilate Synthesis in
Clostridium butyricum
Authors: Baskerville Eugene , Twarog Robert
Published: January 2020

The anthranilate synthetase of
Clostridium butyricum
is composed of two nonidentical subunits of unequal size. An enzyme complex consisting of both subunits is required for glutamine utilization in the formation of anthranilic acid. Formation of anthranilate will proceed in the presence of partially pure subunit I provided ammonia is available in place of glutamine. Partially pure subunit II neither catalyzes the formation of anthranilate nor possesses anthranilate-5-phosphoribosylpyrophosphate phosphoribosyltransferase activity. The enzyme complex is stabilized by high subunit concentrations and by the presence of glutamine. High KCl concentrations promote dissociation of the enzyme into its component subunits. The synthesis of subunits I and II is coordinately controlled with the synthesis of the enzymes mediating reactions 4 and 5 of the tryptophan pathway. When using gel filtration procedures, the molecular weights of the large (I) and small (II) subunits were estimated to be 127,000 and 15,000, respectively. Partially pure anthranilate synthetase subunits were obtained from two spontaneous mutants resistant to growth inhibition by 5-methyltryptophan. One mutant, strain mtr-8, possessed an anthranilate synthetase that was resistant to feedback inhibition by tryptophan and by three tryptophan analogues: 5-methyl-tryptophan, 4- and 5-fluorotryptophan. Reconstruction experiments carried out by using partially purified enzyme subunits obtained from wild-type, mutant mtr-8 and mutant mtr-4 cells indicate that resistance of the enzyme from mutant mtr-8 to feedback inhibition by tryptophan or its analogues was the result of an alteration in the large (I) subunit. Mutant mtr-8 incorporates [
14
C]tryptophan into cell protein at a rate comparable with wild-type cells. Mutant mtr-4 failed to incorporate significant amounts of [
14
C]tryptophan into cell protein. We conclude that strain mtr-4 is resistant to growth inhibition by 5-methyltryptophan because it fails to transport the analogue into the cell. Although mutant mtr-8 was isolated as a spontaneous mutant having two different properties (altered regulatory properties and an anthranilate synthetase with altered sensitivity to feedback inhibition), we have no direct evidence that this was the result of a single mutational event.
Magnesium Transport in
Bacillus subtilis
W23 During Growth and Sporulation
Authors: Scribner Harvey , Eisenstadt Eric , Silver Simon
Published: January 2020

The active transport of magnesium by cells of
Bacillus subtilis
strain W23 occurs by a highly specific transport system (Mg
2+
is favored over Mn
2+
, Co
2+
, or Ca
2+
) that is energy dependent (i.e., glucose is required in minimal medium and the system is inhibited by cyanide and
m
-chlorophenyl carbonylcyanidehydrazone). The rate of magnesium uptake by log-phase
B. subtilis
cells follows saturation kinetics with a

K
m

of 2.5 × 10
−4
M and a

V
max

of 4.4 μmol per min per g (dry weight) at 30 C. Manganese is a competitive inhibitor showing a

K
i

of 5 × 10
−4
M. During sporulation the rate of magnesium transport declines. This decline in rate is specific for the magnesium system as the manganese and calcium transport rates increase. The residual magnesium transport function in sporulating cells shows both an altered

K
m

and an altered

V
max

. The magnesium content of late sporulating cells is also lower than that for log-phase cells.
Chromosomal Location of the
tolG
Locus for Tolerance to Bacteriocin JF246 in
Escherichia coli
K-12
Authors: Foulds John
Published: January 2020

The
tolG
gene is located between
pyrD
and
fabA
on the
Escherichia coli
chromosome.
Dictyostelium discoideum
: a New Method for Cloning in Liquid Medium
Authors: Anderson James J.
Published: January 2020

Dictyostelium discoideum
can be cloned in droplets of liquid medium by a method that permits observation of the complete life cycle. This alternative to solidified media provides advantages in clonal analysis.
Glucose Transport in
Brucella abortus
Authors: Rest Richard F. , Robertson Donald C.
Published: January 2020

Brucella abortus
British strain 19 transported glucose with an apparent

K
m

of 0.16 mM and an apparent

V
max

of 250 nmol per min per mg of N. The only common glucose analogue transported was 2-deoxyglucose (2-DOG), with an apparent

K
i

of 0.73 mM. Alpha- or beta-methyl glucosides and 3-
O
-methylglucose were not transported. Transport was linear for 70 to 90 s, depending on the concentration of substrate used. 2-Deoxyglucose was transported as the free sugar and was not further metabolized once inside the cell. There was no glucose phosphoenolpyruvate phosphotransferase system (PEP-PTS) present, and there were no inhibitors present in
Brucella
cell-free extract that inhibited the
Escherichia coli
glucose PEP-PTS.
N
-Ethylmaleimide (NEM) and
p
-chloromercuribenzoate (
p
CMB) completely inhibited transport of glucose and 2-DOG. Glutathione, dithiothreitol, and β-mercaptoethanol reversed the effects of
p
CMB but not of NEM. A pH optimum of 7.2 and a temperature optimum of 37 to 45 C were observed for both

K
m

and

V
max

. The glucose transport system appeared to be constitutive for glucose transport in cells grown on fructose, galactose, erythritol, or glucose. The electron transfer inhibitors carbonyl cyanide,
m
-chlorophenylhydrazone, NaN
3
, 2,4-dinitrophenol, and KCN inhibited 2-DOG transport to a greater extent than did the metabolic energy inhibitors NaAsO
4
, iodoacetate, KF, and 2-heptyl-4-hydroxyquinoline-
N
-oxide. Dicyclohexylcarbodiimide, an inhibitor of membrane-bound adenosine triphosphatases, inhibited transport by 100%.
Exclusion of Induced Bacteriophage from Cells of a Lysogenic
Bacillus megaterium
Committed to Sporulation
Authors: Hendry G. S. , Fitz-James P. C.
Published: January 2020

Spontaneous release of the temperate bacteriophage T (φT), carried by
Bacillus megaterium
899a, occurred during early growth of the host cells. Rejuvenated cells (accomplished by a 5× dilution in fresh medium) and unrejuvenated cells were induced by mitomycin C during the course of sporulation and subsequent phage φT production measured by burst size. Induction of sequential samples of unrejuvenated cells resulted in burst sizes that fell to zero as T
0
sporulation time in the main culture was approached. This drop in burst size was not considered a sporulation event, as it also occurred during analogous stages of growth in an asporogenous mutant. Rejuvenated, induced portions of the culture of sporulating cells of
B. megaterium
899a gave large burst sizes until T
+2
, when the burst sizes fell to zero. The stage I asporogenous mutant, treated in a similar manner, gave lower, but still substantial, burst sizes; thus, the sharp decline in burst size of induced rejuvenated sporulating cells appeared to be a sporulation event. Sporulating cells induced at times shortly after T
1.5
formed spores in which the induced phage were trapped until germination of the spores, which formed infectious centers. This induced phage-trapping was maximal when the sporulating cultures were induced at T
2.25
. Commitment to sporulation could be defined by our system as that point beyond which rejuvenated sporulating cells were unable to support the replication of the phage. This point also correlated with the increase in induced phage-trapping by spores. Two other methods gave a similar commitment time. Commitment to sporulation, in spite of added glucose or fresh complex medium, occurred at the same time. Electron micrography showed that the committed cell was still undergoing engulfment. The fate of induced phage φT was determined at different points during growth and sporulation. Induction at times prior to T
0
, which were longer than the eclipse period of the phage, resulted in a burst size of approximately 50. At times prior to T
0
, shorter than the phage eclipse period, induction led to lysis with low burst sizes, approaching zero. The pattern of spontaneous phage release during growth was similar. From T
0
up to the point of commitment to sporulation, induction resulted in the blocking of spore formation without lysis. At the commitment point, induced phage were trapped and carried into spores which germinated to give infectious centers. The spontaneous derepression of phage at a time which blocked spore formation led to 7 × 10
4
infectious centers per ml and would not normally be noticed. Derepression at the time of phage entrapment was not observed to occur without induction with mitomycin C.
Properties and Developmental Roles of the Lysyl- and Tryptophanyl-Transfer Ribonucleic Acid Synthetases of
Bacillus subtilis
: Common Genetic Origin of the Corresponding Spore and Vegetative Enzymes
Authors: Steinberg William
Published: January 2020

The lysyl-transfer ribonucleic acid synthetase (LRS) and tryptophanyl-transfer ribonucleic acid synthetases (TRS) (
l
-lysine:tRNA ligase [AMP], EC 6.1.1.6; and
l
-tryptophan:tRNA ligase [AMP], EC 6.1.1.2) have been purified 60- and 100-fold, respectively, from vegetative cells and spores of
Bacillus subtilis
. There are no significant differences between the corresponding spore and vegetative enzymes with respect to their elution characteristics from columns of phosphocellulose or hydroxylapatite, their molecular weight (~130,000 for LRS and ~87,000 for TRS as determined by gel filtration), their kinetic constants for substrates (in the amino acid-dependent adenosine triphosphate-pyrophosphate exchange reaction), and the kinetics of inactivation by heat and by antibody. The Mg
2+
requirement for optimal enzyme activity of the corresponding spore and vegetative enzyme differ slightly. Mutants having defective (temperature sensitive) vegetative LRS or TRS activities produce spores in which these enzymes are also defective. The mutant spores are more heat sensitive than the parental type, but contain normal levels of dipicolinic acid. They germinate normally at the restrictive temperature (43 C), but are blocked at specific developmental stages in outgrowth. No modification in temperature sensitivity phenotype occurs during outgrowth, nor is there a change in molecular weight of the two enzymes. The implication is that the LRS and TRS activities of the vegetative and spore stages are each coded (at least in part) by the same structural gene. The temperature sensitivity of mutant spores is discussed with respect to those factors which are involved in the formation of the heat-resistant state.
Inactivation of Isocitrate Lyase During Myxospore Development in
Myxococcus xanthus
Authors: Orlowski Michael , White David
Published: January 2020

The inactivation of isocitrate lyase which occurs during late stages of myxospore formation in
Myxococcus xanthus
was studied. Several findings are reported. (i) Protein synthesis is required over a specific time interval in order for isocitrate lyase inactivation to occur at a later time. (ii) Metabolic energy is required at all times during myxospore development if the inactivation is to occur. (iii) It was possible to inhibit protein turnover to a considerable extent without affecting the net loss in isocitrate lyase activity.
Effect of Inhibition of Deoxyribonucleic Acid and Protein Synthesis on the Direction of Cell Wall Growth in
Streptococcus faecalis
Authors: Higgins M. L. , Daneo-Moore L. , Boothby D. , Shockman G. D.
Published: January 2020

Selective inhibition of protein synthesis in
Streptococcus faecalis
(ATCC 9790) was accompanied by a rapid and severe inhibition of cell division and a reduction of enlargement of cellular surface area. Continued synthesis of cell wall polymers resulted in rapid thickening of the wall to an extent not seen in exponential-phase populations. Thus, the normal direction of wall growth was changed from a preferential feeding out of new wall surface to that of thickening existing cell surfaces. However, the overall manner in which the wall thickened, from nascent septa toward polar regions, was the same in both exponential-phase and inhibited populations. In contrast, selective inhibition of deoxyribonucleic acid (DNA) synthesis using mitomycin C was accompanied by an increase in cellular surface area and by division of about 80% of the cells in random populations. Little or no wall thickening was observed until the synthesis of macromolecules other than DNA was impaired and further cell division ceased. Concomitant inhibition of both DNA and protein synthesis inhibited cell division but permitted an increase in average cell volume. In such doubly inhibited cells, walls thickened less than in cells inhibited for protein synthesis only. On the basis of the results obtained, a model for cell surface enlargement and cell division is presented. The model proposes that: (i) each wall enlargement site is influenced by an individual chromosome replication cycle; (ii) during chromosome replication peripheral surface enlargement would be favored over thickening (or septation); (iii) a signal associated with chromosome termination would favor thickening (and septation) at the expense of surface enlargement; and (iv) a factor or signal related to protein synthesis would be required for one or more of the near terminal stages of cell division or cell separation, or both.
Cell Separation in Blue-Green Bacteria
Authors: Ingram L. O. , Aldrich H. C.
Published: January 2020

Autolysin-like enzymes appear to be responsible for cell separation in
Agmenellum quadruplicatum
. Mutants that are impaired in cell separation and grow as chains exhibit reduced cell lytic activity. Lysozyme, extracted autolysin, and antibiotics that affect peptidoglycan synthesis phenotypically suppress chain formation. Various aspects of the regulation of the cell separation process were also examined. Studies involving antibiotic inhibitors of macromolecular synthesis and general growth inhibitors provided no evidence for the active regulation of the cell separation process during the latter portion of the division cycle. Evidence was obtained, however, for the partial restriction of peptidogly-can hydrolysis by unknown secondary modifications. The thin electron-dense layer of peptidoglycan along the sides of cells was much more resistant to hydrolysis by egg-white lysozyme than was the septum between daughter cells. The middle portion of the septum was more sensitive than was the layer immediately adjacent to the cytoplasmic membrane. Under conditions that would not osmotically stabilize spheroplasts, lysozyme facilitates rapid cell separation in chain-forming mutants with little leakage of cellular protein or loss of viability.
Intracellular Localization of Nigeran in the Wall of
Aspergillus aculeatus
by Autoradiography with the Electron Microscope
Authors: Gold Michael H. , Larson Susan , Segel Irwin H. , Stocking C. R.
Published: January 2020

Aspergillus aculeatus
mycelium was incubated with [
3
H]glucose under conditions that promote nigeran (mycodextran) accumulation (low pH, in the absence of nitrogen). Autoradiography revealed that essentially all the
3
H was localized around the hyphal perimeter. The results strongly support a hyphal wall location for nigeran.
Attractant-Directed Motility in
Escherichia coli
: Requirement for a Fluid Lipid Phase
Authors: Lofgren Kelly W. , Fox C. Fred
Published: January 2020

Attractant-directed motility (chemotaxis) in
Escherichia coli
has an absolute requirement for a fluid membrane. No such requirement was detected for motility per se.
Transition of the R Factor R12 in
Proteus mirabilis
Authors: Morris C. F. , Rownd R.
Published: January 2020

When
Proteus mirabilis
harboring the R factor R12 (a round of replication mutant of the R factor
NR1
) is cultured in medium containing streptomycin there can be an amplification in the number of copies of r-determinants per cell and the formation of enlarged polygenic R factors containing repeated sequences of r-determinants as well as polygenic molecules consisting of repeated sequences of r-determinants. This phenomenon has been referred to as the “transition.” When transitioned cells are then cultured in drug-free medium, within a few generations two distinct density species of R factor deoxyribonucleic acid (DNA) are observed in a CsCl density gradient: a 1.712 g/ml band of covalently closed circular R factor DNA consisting of one transfer factor (RTF-TC) plus one r-determinant and a 1.718 g/ml band consisting of repeated sequences of r-determinants. The RTF-TC component of the R factor appears to control the replication of all the R factor DNA which is attached to it. In the autonomous state, however, polygenic sequences of r-determinants do not appear to replicate under the same control mechanism as when they are attached to an RTF-TC.
Septum Formation inEscherichia coli: Characterization of Septal Structure and the Effects of Antibiotics on Cell Division
Authors: Burdett I. D. J. , Murray R. G. E.
Published: January 2020

Septa can be demonstrated in sections ofEscherichia colistrains B and B/r after fixation with acrolein and glutaraldehyde. The septum consists of an ingrowth of the cytoplasmic membrane and the mucopeptide layer; the outer membrane is excluded from the septum until the cells begin to separate. Mesosomes have also been observed. The septum is highly labile and, except in the chain-forming strains,E. coliD22envA and CRT 97, not easily preserved by standard procedures. The labile nature of the septum may be due to the presence of autolysin(s) located at the presumptive division site. Blocking division by addition of ampicillin (2 to 5 μg/ml) to cells ofE. coliB/r produces a bulge at the middle of the cells; bulge formation is stopped by addition of chloramphenicol. Cephalosporins also induce bulge formation but may stop cell elongation as well as division. Bulge formation, due to the presumed action of an autolysin(s), may be an initial step in the septation sequence when the mucopeptide is modified to allow construction of the septum. In a nonseptate filament-forming strain, PAT 84, which ceases to divide at 42 C, bulge formation only occurs in the presence of ampicillin at the time of a shift-down at 30 C or at 42 C in the presence of NaCl (0.25 to 0.34 M). Experiments with chloramphenicol suggest that the filaments are fully compartmentalized but fail to divide owing to the inactivation, rather than loss of synthesis, of an autolysin at 42 C.
Peptide Utilization by Amino Acid Auxotrophs of
Neurospora crassa
Authors: Wolfinbarger L. , Marzluf George A.
Published: January 2020

The ability of auxotrophs of
Neurospora crassa
to grow on certain tripeptides, despite the presence of excess competing amino acids, suggests it has an oligopeptide transport system. In general, dipeptides did not support growth except in those instances where extracellular hydrolysis occurred, or where the dipeptide appeared to be accumulated by an uptake system which is sensitive to inhibition by free amino acids. Considerable intracellular peptidase activity toward a large number of peptides was demonstrated, including a number of peptides which could not be utilized for growth. The intracellular peptidase activity was shown to be selective for amino acid composition and sequence (N-terminal or C-terminal) within the peptide; glycine-containing peptides were particularly poor substrates for peptidase activity. Only a small amount of extracellular peptidase activity could be detected.
Clustering of the Genes for Allantoin Degradation in
Saccharomyces cerevisiae
Authors: Lawther Robert P. , Riemer Elaine , Chojnacki Bonnie , Cooper Terrance G.
Published: January 2020

We have shown that allantoin degradation in
Saccharomyces cerevisiae
proceeds exclusively through the intermediate formation of allantoic acid, urea, and allophanic acid. The number of reactions between allantoic acid and urea, however, remains obscure owing to our inability to isolate a mutant defective in ureidoglycolate hydrolase. Structural genes for the enzymes, allantoinase (
dal1
) and allantoicase (
dal2
) are located on chromosome IX promixal to the centromere in the order
dal1-dal2-lysl
.
Time Scale for Rejoining of Bacteriophage λ Deoxyribonucleic Acid Molecules in Superinfected
pol
+
and
polA1
Strains of
Escherichia coli
After Exposure to 4 MeV Electrons
Authors: Boye Erik , Johansen Ivar , Brustad Tor
Published: January 2020

The time scale for rejoining of radiation-induced deoxyribonucleic acid (DNA) single-strand breaks was measured in the presence and absence of oxygen. The involvement of DNA polymerase I in this repair process was studied. Formation and rejoining of DNA strand breaks were measured in λ DNA infecting lysogenic
pol
+
and
polA1
strains of
Escherichia coli
irradiated by 4 MeV electrons under identical conditions. Irradiation and transfer to alkaline detergent could be completed in less than 180 ms. The initial yields of DNA strand breaks were identical in
pol
+
and
polA1
host cells and four- to fivefold higher in the presence of oxygen than in nitrogen anoxia. Evidence for the existence of a very fast repair process, independent of DNA polymerase I, was not found, since no rejoining of radiation-induced DNA strand breaks was observed during incubation from 45 ms to 3 s. In
pol
+
host cells most of the strand breaks produced in the presence of oxygen were rejoined within the first 30 to 40 s of incubation, whereas no rejoining could be detected within the same period of time in anoxic cells. Since no rejoining of broken λ DNA molecules was observed in
polA1
host cells, it is concluded that the synthetase activity of DNA polymerase I is involved in the rejoining of DNA breaks induced by radiation in the presence of oxygen.
Attempts to Detect Deoxyribonucleic Acid from
Agrobacterium tumefaciens
and Bacteriophage PS8 in Crown Gall Tumors by Complementary Ribonucleic Acid/Deoxyribonucleic Acid-Filter Hybridization
Authors: Eden Francine C. , Farrand Stephen K. , Powell Jerry S. , Bendich Arnold J. , Chilton Mary-Dell , Nester Eugene W. , Gordon Milton P.
Published: January 2020

Labeled ribonucleic acid (RNA) complementary to
Agrobacterium tumefaciens
DNA and PS8 bacteriophage DNA (cRNA) were used in a systematic study of the sensitivity of cRNA/deoxyribonucleic acid (DNA)-filter hybridization for detection of small amounts of phage or bacterial DNA immobilized on filters.
A. tumefaciens
cRNA of specific activity 10
6
to 2 × 10
6
counts per min per μg reacted to a significant extent when the DNA-filter contained 1%
A. tumefaciens
DNA in a salmon DNA background, but 0.1%
A. tumefaciens
DNA was not detectable. PS8 phage cRNA of the same specific activity reacted to a significant extent when the DNA-filter contained as little as 0.01% PS8 DNA in a salmon DNA background. Both kinds of cRNA were found to bind to tobacco crown gall tumor DNA-filters. Similar reaction was found with control normal callus DNA-filters but not with tobacco seedling DNA-filters. The “hybrids” formed by cRNA with normal callus and tumor DNA-filters had low thermal stability. Attempts to purify the tumor and normal callus DNA prior to immobilization on the filter resulted in elimination of this spurious binding. No evidence was found for bacterial or phage DNA in crown gall tumor DNA.