bluuebunny/crossref_metadata_2025 huggingface dataset:
Phenotypic Recognition of Pyocyanine Mutants in
Pseudomonas aeruginosa
Authors: Carson Melinda , Jensen Roy A.
Published: January 2020

Certain classes of pyocyanine mutants in
Pseudomonas aeruginosa
escape detection when screened in the presence of wild-type or other mutant cells. A technique is described for recognizing mutant phenotypes after cells are in individual agar wells. The procedure eliminates cross-feeding phenomena as well as the masking of mutant phenotypes by diffusing pyocyanine produced by nearby clones.
Sporulation of
Bacillus thuringiensis
Without Concurrent Derepression of the Tricarboxylic Acid Cycle
Authors: Nickerson Kenneth W. , De Pinto John , Bulla Lee A.
Published: January 2020

Bacillus thuringiensis
sporulates in a glucose-glutamate medium without concurrent derepression of the tricarboxylic acid cycle. Glutamate appears to regulate tricarboxylic acid cycle activity as well as to influence spore heat resistance and production of dipicolinic acid.
Characterization of the Antigenic Subunits of the Envelope Protein ofYersinia pestis
Authors: Bennett Larry G. , Tornabene Thomas G.
Published: January 2020

Chemical, physical, and immunological properties of the envelope antigen ofYersinia pestisstrains have been investigated. The antigen consists of two components with isoelectric points (pI) of 4.6 and 4.8. One component (pI 4.6) is a protein bound to a small carbohydrate moiety identified as an oligomeric galactan; the other component (pI 4.8) is a simple protein. These two components are antigenically identical. In buffered solution, the antigen exists as aggregates of molecular weights larger than 300,000. The aggregates dissociate into a variety of smaller molecular weight forms depending on the nature of the treatment for dissociation. Each aggregate can be further dissociated into a single antigenic subunit fraction containing protein and glycoprotein species with molecular weights in the range from 15,000 to 17,000. The subunits can be obtained by a dissociation treatment with 0.1% mercaptoethanol in 0.25% sodium dodecyl sulfate at 95 C for 5 min. The subunits will readily reaggregate into a variety of larger molecular weight forms on the removal of dodecyl sulfate.
Germination of Yeast Spores Lacking Mitochondrial Deoxyribonucleic Acid
Authors: Tingle Marjorie A. , Küenzi Martin T. , Halvorson Harlyn O.
Published: January 2020

A population of petite ascospores (mitochondrial deoxyribonucleic acid [mtDNA]-less), produced by brief ethidium bromide (EthBr) mutagenesis prior to transfer to sporulation medium, was used to examine the role of the mitochondrial genetic system on germination and outgrowth in
Saccharomyces cerevisiae
. Petite ascospores, which are morphologically indistinguishable by phase-contrast microscopy from wild-type spores, germinate and proceed through outgrowth at a rate and extent only slightly less than that of wild-type spores. Both developmental processes occurred in the absence of mtDNA synthesis and measurable cytochrome oxidase activity. These results indicate that neither respiration nor a functional mitochondrial genome are required for germination and outgrowth. The properties of the petite clones were typical of petites formed during vegetative growth. Individual sporal clones differed markedly from each other in suppressiveness. Petite sporal clones which exhibited a high degree of supressiveness also contained a reduced but detectable amount of mtDNA of altered buoyant density. One clone contained a unique mtDNA with a buoyant density higher than that of wild-type mtDNA.
Localization of Two Functions of the Phosphoribosyl Anthranilate Transferase of
Escherichia coli
to Distinct Regions of the Polypeptide Chain
Authors: Jackson Ethel Noland , Yanofsky Charles
Published: January 2020

The
trpD
gene specifies a polypeptide which has both glutamine amidotransferase and phosphoribosyl anthranilate (PRA) transferase activities. Deletions fusing segments of
trpD
to the gene preceding it in the operon,
trpE
, were selected in strains carrying various
trpD
point mutations. The selection procedure required both that a deletion enter
trpE
and that it restore the PRA transferase activity which the parent
trpD
point mutant lacked. Deletion mutants were found which had PRA transferase activity although the first third of
trpD
was deleted. The existence of the mutants proves that a terminal segment of
trpD
is sufficient to specify a polypeptide having PRA transferase activity. The location of the deletion end points on the genetic map of
trpD
defines the extent of the
trpD
segment required for PRA transferase activity. This segment did not overlap the initial region of
trpD
required to specify the glutamine amidotransferase function of the
trpD
polypeptide. These results support the hypothesis (M. Grieshaber and R. Bauerle, 1972; H. Zalkin and L. H. Hwang, 1971) that the bifunctional
trpD
polypeptide might have evolved by fusion of a gene specifying a glutamine amidotransferase with a gene directing PRA transferase synthesis.
Penicillin-Resistant Temperature-Sensitive Mutants of
Escherichia coli
Which Synthesize Hypo- or Hyper-Cross-Linked Peptidoglycan
Authors: Kamiryo Tatsuyuki , Strominger Jack L.
Published: January 2020

A group of
Escherichia coli
mutants which are ampicillin resistant at 32 C and which either are unable to grow or lyse at 42 C has been selected. These mutants have been classified by a number of characteristics: total peptidoglycan synthesis measured by [
14
C]diaminopimelic acid incorporation, extent of cross-linking of the peptidoglycan which is synthesized, growth characteristics at the two temperatures, and morphology. Two especially interesting groups of mutants have been described. In one of these, a hypo-cross-linked peptidoglycan was synthesized at the nonpermissive temperature. Most of these organisms lysed at 42 C. In another group, the peptidoglycan synthesized at 42 C was hyper-cross-linked. Many of these organisms were spherical. Studies of revertants indicated that ampicillin resistance, temperature sensitivity, cross-linking, growth characteristics, and morphological changes may be related to a single mutational event in both of these groups.
Mechanism of Action and Development of Resistance to a New Amidino Penicillin
Authors: Matsuhashi Sachiko , Kamiryo Tatsuyuki , Blumberg Peter M. , Linnett Paul , Willoughby Eileen , Strominger Jack L.
Published: January 2020

The mechanism of killing of
Escherichia coli
by a novel beta-lactam antibiotic, an amidino penicillin, has been investigated. This compound converts
E. coli
to relatively stable spherical forms at low concentration. However, the amidino penicillin caused no alteration in any of those parameters of peptidoglycan synthesis which can be studied. Above 10 μg of the antibiotic per ml the cells began to lyse, and a second mode of killing appeared. Mutants resistant to the amidino penicillin were isolated and several were studied in detail. Three mutant phenotypes were distinguished: (i) spherical shape and hypersensitive to lysis by either amidino penicillin or ampicillin; (ii) spherical shape and normally sensitive to lysis; (iii) rod shape, converted to viable spheres by amidino penicillin and normally sensitive to lysis.
Histidyl-Transfer Ribonucleic Acid Synthetase in Positive Control of the Histidine Operon in
Salmonella typhimurium
Authors: Wyche James H. , Ely Bert , Cebula Thomas A. , Snead Michael C. , Hartman Philip E.
Published: January 2020

Histidyl-transfer ribonucleic acid (tRNA) synthetase (HRS), coded by the
hisS
gene, appears to play two roles in regulation of the histidine operon of
Salmonella typhimurium
: (i) in synthesis of a critical effector molecule, histidyl-tRNA, and (ii) a more direct effect elicited by the presence of the enzyme protein itself. The specific activity of HRS was elevated either by mutations in the
strB
locus or in
hisS
+
merodiploids of
Escherichia coli/S. typhimurium
and
S. abony/S. typhimurium
. In each case, an increase in HRS was accompanied by an increase in histidine operon expression, indicating that HRS may be involved in positive control of the histidine operon. It is unlikely that HRS leads to increased histidine operon expression merely by acting as a “sponge” for charged tRNA. Rather, HRS appears to influence operon expression by interaction with some effector molecule other than charged tRNA or by a direct interaction with the histidine operator-promoter region. The functional level of histidine operon expression has no effect on HRS specific activity.
Inhibition of the
Bacillus subtilis
Membrane-Bound
d
-Alanine Carboxypeptidase by 6-Aminopenicillanic Acid Covalently Coupled to Sepharose
Authors: Storm Daniel R. , Blumberg Peter M. , Strominger Jack L.
Published: January 2020

An insoluble penicillin derivative, 6-aminopenicillanic acid covalently coupled to Sepharose, was used to investigate the location of the
d
-alanine carboxypeptidase in the
Bacillus subtilis
membrane. Only 50% of the enzymatic activity in protoplasts and 74% of that in purified membranes was sensitive, although the purified enzyme could be inhibited completely by the resin. These results suggest that a minimum of 50% of the
d
-alanine carboxypeptidase exists on the outer face of the bacterial membrane.
Regulation of Tryptophan Biosynthesis in
Saccharomyces cerevisiae
: Mode of Action of 5-Methyl-Tryptophan and 5-Methyl-Tryptophan-Sensitive Mutants
Authors: Schürch A. , Miozzari J. , Hütter R.
Published: January 2020

In a wild-type strain of
Saccharomyces cerevisiae
the tryptophan analogue
dl
-5-methyl-tryptophan (5MT) causes only a slight reduction of the growth rate. Uptake experiments indicate that the limited inhibition is partly due to low levels of 5MT inside the cell. On the other hand, this low concentration of 5MT leads to an increase in the activity of the tryptophan-biosynthetic enzymes. Evidence is presented that suggests that 5MT acts primarily through feedback inhibition of anthranilate synthase, the first enzyme of the pathway. A number of 5MT-sensitive mutants have been isolated, characterized, and assigned to one of the following three classes: class I, strains with altered activity and/or feedback sensitivity of anthranilate synthase; class II, strains with elevated uptake of 5MT; class III, mutants with altered regulation of the tryptophan-biosynthetic enzymes, which do not exhibit increases in activity in the presence of 5MT. This failure to exhibit increased enzyme activities in mutants of class III can also be observed after tryptophan starvation. Two mutants of class III show high sensitivity towards 3-amino-1,2,4-triazole. They can not exhibit derepression of some histidine- and arginine-biosynthetic enzymes under conditions that lead to an increase in these same enzymes in the wild-type strain.
Size Distribution of the Closed Circular Deoxyribonucleic Acid Molecules of
Bacillus megaterium
: Sedimentation Velocity and Electron Microscope Measurements
Authors: Carlton Bruce C. , Smith Malcolm P. W.
Published: January 2020

The circular deoxyribonucleic acid (DNA) of
Bacillus megaterium
was fractionated by sedimentation velocity on preparative zonal gradients. The fractions obtained were characterized by analytical sedimentation velocity analysis on neutral and alkaline sucrose gradients, and by contour length measurement by electron microscopy. Each fraction was found to contain circular molecules of one to three discrete sizes, in various combinations of covalently closed and open circular forms. Estimations of the molecular masses of these sizes gave values of 3.9, 6.2, 16.0, 31, and 60 million daltons for the major molecular species. Minor amounts of molecules of 7.6, 11.7, 47, 89, and 112 million daltons were observed in the electron microscope analyses. Length measurements of almost 600 molecules from the various fractions showed that all except six could be placed in distinct size classes. The distribution of molecular sizes in unfractionated circular DNA was shown to consist primarily of the two smallest size classes, although the relative proportions by weight of five of the classes were roughly equivalent.
Transport of Glucose, Gluconate, and Methyl α-D-Glucoside by
Pseudomonas aeruginosa
Authors: Guymon L. F. , Eagon R. G.
Published: January 2020

Glucose transport by
Pseudomonas aeruginosa
was studied. These studies were enhanced by the use of a mutant, strain PAO 57, which was unable to grow on glucose but which formed the inducible glucose transport system when grown in media containing glucose or other inducers such as 2-deoxy-
d
-glucose. Both PAO 57 and parental strain PAO transported glucose with an apparent

K
m

of 7 μM. Free glucose was concentrated intracellularly by
P. aeruginosa
PAO 57 over 200-fold above the external level. These data constitute direct evidence that glucose is transported via active transport by
P. aeruginosa
. Various experimental data clearly indicated that
P. aeruginosa
PAO transported methyl α-
d
-glucose (α-MeGlc) via the glucose transport system. The apparent

K
m

of α-MeGlc transport was 7 mM which indicated a 1,000-fold lower affinity of the glucose transport system for α-MeGlc than for glucose. While only unchanged α-MeGlc was detected intracellularly in
P. aeruginosa
, α-MeGlc was actually concentrated intracellularly less than 2-fold over the external level. Membrane vesicles of
P. aeruginosa
PAO retained transport activity for gluconate. This solute was concentrated intravesicularly several-fold over the external level. A component of the glucose transport system is believed to have been lost during vesicle preparation since glucose per se was not transported. Instead; glucose was converted to gluconate by membrane-associated glucose dehydrogenase and gluconate was then transported into the vesicles. Although this may constitute an alternate system for glucose transport, it is not a necessary prerequisite for glucose transport by intact cells since
P. aeruginosa
PAO 57, which lacks glucose dehydrogenase, was able to transport glucose at a rate equal to the parental strain.
Retraction of F Pili
Authors: Novotny Charles P. , Fives-Taylor Paula
Published: January 2020

The disappearance of F pili on
Escherichia coli
cells in the presence of 10
−2
M NaCN was studied by electron microscopy and serum-blocking power. The pili which disappeared from the cell did not appear as free pili in the culture medium, suggesting that the pili had retracted into the cell. New pili were produced at a normal rate approximately 3 min after NaCN was removed. The adsorption of either F pili antibody or R17 bacteriophage to the sides of pili and temperatures below 24 C prevented retraction. The disappearance of pili was accompanied by a loss in the ability of cells to adsorb R17 phage and the type of F pili antibody that inhibits R17 phage infection and mating. The ability to adsorb M13 phage and the type of F pili antibody that inhibits M13 phage infection was not affected. This suggests that the tips of retracted pili are exposed.
Proteins of the Inner Membrane of
Escherichia coli
: Changes in Composition Associated with Anaerobic Growth and Fumarate Reductase Amber Mutation
Authors: Spencer Margaret E. , Guest J. R.
Published: January 2020

The inner membrane fractions of
Escherichia coli
grown anaerobically and aerobically were isolated, and their proteins were compared by electrophoresis in polyacrylamide gels. To maximimize the differences between the preparations, the anaerobic cultures were grown on complex medium with added glucose, but glucose was omitted from the aerobic cultures to prevent catabolite repression. The pattern of bands in the two types of preparation differed considerably, and changes in approximately 20 components were observed. In particular, the band identified as succinate dehydrogenase in aerobic preparations was greatly reduced in anaerobic preparations. Mutants lacking fumarate reductase were isolated, and inner membrane preparations of an
frd
amber mutant were deficient in a major component of 75,000 daltons and possibly a minor one of 87,500 daltons. The former was also present in greater amounts in anaerobic preparations and could represent a fumarate reductase subunit.
Purification and Properties of Dextransucrase from
Streptococcus mutans
Authors: Chludzinski Andrew M. , Germaine Greg R. , Schachtele Charles F.
Published: January 2020

The dextransucrase (EC 2.4.1.5) activity from cell-free culture supernatants of
Streptococcus mutans
strain 6715 has been purified approximately 1,500-fold by ammonium sulfate precipitation, hydroxylapatite chromatography, and isoelectric focusing. The enzyme was eluted as a single peak of activity from hydroxylapatite, and isoelectric focusing of the resulting preparation gave a single band of dextransucrase activity which focused at a pH of 4.0. The final enzyme preparation contained two distinct, enzymatically active proteins as judged by assay in situ after polyacrylamide gel electrophoresis. One of the proteins represented 90% of the total dextransucrase activity and 53% of the total protein. The molecular weight of the enzyme was estimated by gel filtration to be 94,000. The temperature optimum of the enzyme was broad (34 to 42 C) and its pH range was rather narrow, with optimal activity at pH 5.5. The

K
m

for sucrose was 3 mM, and fructose competitively inhibited the enzyme reaction with a

K
i

of 27 mM.
Glutamate Synthase: Properties of the Reduced Nicotinamide Adenine Dinucleotide-Dependent Enzyme from
Saccharomyces cerevisiae
Authors: Roon Robert J. , Even Harvey L. , Larimore Fred
Published: January 2020

A reduced nicotinamide adenine dinucleotide (NADH)-dependent glutamate synthase has been detected and partially purified from crude extracts of
Saccharomyces cerevisiae
. The enzyme is specific for NADH, glutamine, and α-ketoglutarate (

K
m

values of 2.6 μM, 1.0 mM, and 140 μM, respectively) and has a pH optimum between 7.1 and 7.7. The stoichiometry of the reaction has been determined as 2 mol of glutamate synthesized per mol of glutamine consumed. Glutamate synthase can be distinguished from either of the glutamate dehydrogenases of yeast on the basis of its substrate requirements and behavior during agarose gel and ion exchange chromatography. Variations in the specific activity of glutamate synthase, which occur in response to changes in the growth medium, are similar in character to those observed with the nicotinamide adenine dinucleotide phosphate-dependent (anabolic) glutamate dehydrogenase.
Electron Microscope and Autoradiographic Study of Ultrastructural Aspects of Competence and Deoxyribonucleic Acid Absorption in
Bacillus subtilis
: Localization of Uptake and of Transport of Transforming Deoxyribonucleic Acid in Competent Cells
Authors: Vermeulen C. A. , Venema G.
Published: January 2020

With the aid of serial-section electron microscopy two types of mesosomes can be distinguished in cells of competent cultures of
Bacillus subtilis
: (i) mesosomes connected to the plasma membrane only (plasma membrane mesosomes) and (ii) mesosomes which extend from the plasma membrane into the nuclear bodies (nuclear mesosomes). Contrary to plasma membrane mesosomes, nuclear mesosomes are absent from the tip zones. Electron microscopic autoradiography of sections of
Bacillus subtilis
cells exposed to [
3
H]thymidine-labeled transforming deoxyribonucleic acid (DNA) for a short period of time shows that the DNA becomes associated with mesosomes. As a function of time the DNA migrates towards the nucleoids. Transport of DNA is completed within 15 to 60 min after termination of DNA uptake. During its migration the DNA continues to be associated with mesosomes, presumably with nuclear mesosomes. DNA initially associated with plasma membrane mesosomes of the tip zones is probably transported first towards the middle zones peripherally and from there towards the nucleoids.
Regulation of Substrate Uptake in
Nocardia erythropolis
Authors: Pang Roy H. L. , Bradley S. G.
Published: January 2020

The regulation of uptake of glucose (GLU), glycerol (GLY), mannitol (MTL), and succinate (SUC) has been examined in
Nocardia erythropolis
305. The apparent

K
m

values of the uptake activities of cells subcultured in a medium with the corresponding substrate as the sole carbon source were 205, 48, 8.7, and 36 μM for GLU, GLY, MTL, and SUC, respectively. GLU and GLY uptake activities were constitutive, although there was evidence for an additional inducible component in GLY uptake. Moreover, MTL and SUC uptake activities were inducible. MTL uptake activity was markedly induced by cultivation in MTL medium and partially induced by growth in GLU medium, whereas SUC uptake was induced only by cultivation in SUC medium. SUC added to MTL medium partially repressed the formation of, or inhibited the activity of, MTL uptake. When not induced, uptake of MTL and SUC was proportional to the substrate concentration. The induced uptake of MTL and SUC and the constitutive uptake of GLU were energy dependent and carrier mediated. Uptake of GLY, constitutively or when induced, was also carrier mediated.
Porosity of the Yeast Cell Wall and Membrane
Authors: Scherrer Rene , Louden Louise , Gerhardt Philipp
Published: January 2020

The limiting sizes of molecules that can permeate the intact cell wall and protoplast membrane of
Saccharomyces cerevisiae
were determined from the inflection points in a triphasic pattern of passive equilibrium uptake values obtained with a series of inert probing molecules varying in molecular size. In the phase identified with the yeast protoplast, the uptake-exclusion threshold corresponded to a monodisperse ethylene glycol of molecular weight = 110 and Einstein-Stokes hydrodynamic radius (

r
ES

) = 0.42 nm. In the cell wall phase, the threshold corresponded to a polydisperse polyethylene glycol of number-average molecular weight (

¯M
n

) = 620 and average radius (

r
ES

) = 0.81 nm. The third phase corresponded to complete exclusion of larger molecules. The assessment of cell wall porosity was confirmed by use of a second method involving analytical gel chromatographic analyses of the molecular weight distribution for a single polydisperse polyglycol before and after uptake by the cells, which indicated a quasi-monodisperse threshold for the cell wall of

M
n

= 760 and

r
ES

= 0.89 nm. The results were reconciled with two situations in which much larger protein molecules previously have been reported able to penetrate the yeast cell wall.
Regulation of Alanine Dehydrogenase in
Bacillus licheniformis
Authors: McCowen S. M. , Phibbs P. V.
Published: January 2020

Cell extracts of
Bacillus licheniformis
were found to contain nicotinamide adenine dinucleotide (NAD)-dependent
l
-alanine dehydrogenase (ADH) (
l
-alanine: NAD oxidoreductase, EC 1.4.1.1). High specific activities (3.5 to 6.0 IU/mg of protein) were found in extracts of cells throughout growth cycles only when
l
-alanine served as the primary source of carbon or carbon and nitrogen. Specific activities were minimal (0.02 to 0.04 IU/mg of protein) during growth on glucose, but increased at least sevenfold during the first 5 h of postlogarithmic-phase metabolism. Addition of 10 mM glucose to cultures during logarithmic-phase growth on
l
-alanine resulted in a rapid decrease in enzyme activity. Addition of 20 mM
l
-alanine to cells near the completion of log-phase growth on glucose resulted in a 20-fold increase in ADH specific activity during less than one cell generation. Extracts of postlogarithmic-phase cells cultured on glucose, malate,
l
-glutamate, or Casamino Acids contained intermediate levels of ADH activity. The enzyme was partially purified from crude extracts of
B. licheniformis
, and apparent kinetic constants were estimated. A role for ADH in the catabolism of
l
-alanine to pyruvate during vegetative growth on
l
-alanine and during sporulation of cells cultured on glucose is proposed on the basis of these experimental results.
Mapping of the Locus for
Escherichia coli
Transfer Ribonucleic Acid Nucleotidyltransferase
Authors: Foulds John , Hilderman Richard H. , Deutscher Murray P.
Published: January 2020

cca
Mutants of
Escherichia coli
K12 have altered levels of tRNA nucleotidyl-transferase activity. The
cca
locus has been located at minute 59.4 of the
E. coli
linkage map. It is cotransduced with
tolC
but not with
argG
, and is the earliest known marker transferred by Hfr strain KL14. The proximity of the
tolC
locus to the integrated sex factor in Hfr strain KL14 may be useful for mapping sex factor mutations by transduction.
Preparation of Cells Permeable to Macromolecules by Treatment with Toluene: Studies of Transfer Ribonucleic Acid Nucleotidyltransferase
Authors: Deutscher Murray P.
Published: January 2020

Transfer ribonucleic acid (tRNA) nucleotidyltransferase was studied after making cells permeable to macromolecules by treatment with toluene. The conditions of toluene treatment necessary for obtaining maximal activity were defined. Toluene treatment was most efficient when carried out for 5 min at 37 C at pH 9.0 on log-phase cells. No activity could be detected if cells were treated at 0 C, or in the presence of MgCl
2
, or if the cells were in the stationary phase of growth. However, inclusion of lysozyme and ethylenediaminetetraacetic acid during the toluene treatment did render stationary phase cells permeable. The properties of tRNA nucleotidyltransferase from toluene-treated cells were essentially identical to those of purified enzyme with regard to pH optimum, specificity for nucleoside triphosphates and tRNA, and apparent

K
m

values for substrates. In addition to tRNA nucleotidyltransferase, a variety of other enzymes which incorporate adenosine 5′-triphosphate into acid-precipitable material could also be detected in toluene-treated cells. Centrifugation of cells treated with toluene revealed that tRNA nucleotidyltransferase leaked out of cells, whereas other activities remained associated with the cell pellets. Chromatography of the material extracted from toluene-treated cells on Sephadex G-100 indicated that toluene treatment selectively extracts lower molecular weight proteins. The usefulness of such a procedure as an initial step in purification of such enzymes, and its application to tRNA nucleotidyltransferase, is discussed.
Effect of Substrate on the Fatty Acid Composition of Hydrocarbon-Utilizing Filamentous Fungi
Authors: Cerniglia C. E. , Perry J. J.
Published: January 2020

The fatty acid pattern in hydrocarbon-utilizing filamentous fungi was determined after growth on acetate, propionate,
n
-alkanes (C
13
to C
15
), and alk-1-enes (C
14
to C
18
). The fatty acid profile of
Cunninghamella elegans
and
Penicillium zonatum
after growth on acetate shows a predominance of even-carbon fatty acids (C
16
, C
18:1
, C
18:2
), whereas cells grown on propionate showed significantly higher levels of odd-carbon fatty acids (C
15
, C
17
, C
17:1
). Growth on
n
-alkanes resulted in the incorporation of fatty acids homologous to the growth substrate.
Cunninghamella elegans
grown on the alk-1-enes from C
14
to C
18
incorporated the unsaturated substrate into cellular fatty acid after oxidation at the saturated end of the molecule. Regardless of substrate these fungi contain, predominantly, fatty acids 18 carbons in length.
Transport ofd-Arabinose-5-Phosphate andd-Sedoheptulose-7-Phosphate by the Hexose Phosphate Transport System ofSalmonella typhimurium
Authors: Eidels Leon , Rick Paul D. , Stimler Norma P. , Osborn M. J.
Published: January 2020

d-Arabinose-5-phosphate andd-sedoheptulose-7-phosphate were found to be substrates, although not inducers, of the hexose phosphate transport system ofSalmonella typhimurium. Transport of these two sugar phosphates by wild-type strains required preinduction of the hexose phosphate transport system. A mutant ofS. typhimuriumconstitutive for this system also transportedd-arabinose-5-phosphate andd-sedoheptulose-7-phosphate in a constitutive fashion. Glucose-6-phosphate was a potent competitor of the transport of bothd-arabinose-5-phosphate andd-sedoheptulose-7-phosphate. TheKmvalues for transport ofd-glucose-6-phosphate,d-arabinose-5-phosphate, andd-sedoheptulose-7-phosphate were 0.13, 0.32 and 1.61 mM, respectively. The apparentVmaxvalues for transport ofd-glucose-6-phosphate,d-arabinose-5-phosphate, andd-sedoheptulose-7-phosphate were 6.3, 13.2 and 3.0 nmol per min per 5 × 108bacteria, respectively.d-Ribulose-5-phosphate andd-xylulose-5-phosphate did not inhibit transport of the above substrates, whereasd-ribose-5-phosphate was a weak inhibitor ofd-sedoheptulose-7-phosphate transport.
Lysis of Yeast Cell Walls: Glucanases from
Bacillus circulans
WL-12
Authors: Fleet Graham H. , Phaff Herman J.
Published: January 2020

Endo-β-(1 → 3)- and endo-β-(1 → 6)-glucanases are produced in high concentration in the culture fluid of
Bacillus circulans
WL-12 when grown in a mineral medium with bakers' yeast cell walls as the sole carbon source. Much lower enzyme levels were found when laminarin, pustulan, or mannitol was the substrate. The two enzyme activities were well separated during Sephadex G-100 chromatography. The endo-β-(1 → 3)-glucanase was further purified by diethylaminoethyl-cellulose and hydroxyapatite chromatography, whereas the endo-β-(1 → 6)-glucanase could be purified further by diethylamino-ethyl-cellulose and carboxymethyl cellulose chromatography. The endo-β-(1 → 3)-glucanase was specific for the β-(1 → 3)-glucosidic bond, but it did not hydrolyze laminaribiose; laminaritriose was split very slowly. β-(1 → 4)-Bonds in oat glucan in which the glucosyl moiety is substituted in the 3-position were also cleaved. The kinetics of laminarin hydrolysis (optimum pH 5.0) were complex but appeared to follow Michaelis-Menten theory, especially at the lower substrate concentrations. Glucono-δ-lactone was a noncompetitive inhibitor and Hg
2+
inhibited strongly. The enzyme has no metal ion requirements or essential sulfhydryl groups. The purified β-(1 → 6)-glucanase has an optimum pH of 5.5, and its properties were studied in less detail. In contrast to the crude culture fluid, the two purified β-glucanases have only a very limited hydrolytic action on cell wall of either bakers' yeast or of
Schizosaccharomyces pombe
. Although our previous work had assumed that the two glucanases studied here are responsible for cell wall lysis, it now appears that the culture fluid contains in addition a specific lytic enzyme which is eliminated during the extensive purification process.
Homologous and Hybrid Complexes of Anthranilate Synthase from
Bacillus
Species
Authors: Patel Nanu , Holmes W. M. , Kane J. F.
Published: January 2020

The subunits of anthranilate synthase were separated and partially purified by Sephadex G-100 gel filtration from the following six species of
Bacillus
:
Bacillus subtilis, Bacillus licheniformis, Bacillus alvei, Bacillus coagulans, Bacillus pumilus
, and
Bacillus mascerans
. Our data suggest that the enzyme from
B. alvei
is unique among these species. First, the anthranilate synthase complexes are readily dissociated during gel filtration in the absence of glutamine into a large component (aminotransferase), subunit E, and a small component subunit X (glutamine-binding protein), whereas a higher salt concentration is required to dissociate the complex from
B. alvei
. Second, the aminotransferase activity from all six species is stimulated by glycerol and inhibited by tryptophan; however, only the large component from
B. alvei
is stimulated by 2-mercaptoethanol. Finally, the large component can be titrated with the small component to yield a complex which can utilize glutamine as a substrate (amidotransferase). The homologous complexes have an amidotransferase to aminotransferase ratio of 1.4 to 2.3, but the
B. alvei
complex has a ratio of 0.9. Except for complexes that involve the large component from
B. alvei
, hybrid complexes can be formed which have ratios as good as the homologous complexes. These data are consistent with the hypothesis that
B. alvei
is unique among the bacilli with respect to some enzymes in the aromatic amino acid biosynthetic pathway.
Regulation of Nitrogenase Synthesis by Oxygen in
Klebsiella pneumoniae
Authors: St. John Richard T. , Shah Vinod K. , Brill Winston J.
Published: January 2020

Klebsiella pneumoniae
does not fix N
2
under aerobic conditions. The two protein components required for nitrogenase activity were studied during aeration of cells in nitrogen-free media. Component II of nitrogenase was inactivated more slowly in vivo than component I during aeration. The rate of loss of component II was less than the rate of component II synthesis during derepression. No inactive components were detected in cells that had been growing on NH
4
+
and then aerated in nitrogen-free medium. This supports the hypothesis that O
2
somehow represses the formation of nitrogenase.
Regulation of Glycerol Catabolism in
Klebsiella aerogenes
Authors: Ruch F. E. , Lengeler J. , Lin E. C. C.
Published: January 2020

The utilization of glycerol as a carbon source for growth by
Klebsiella aerogenes
, strain 2103, involves separate aerobic (
sn
-glycerol-3-phosphate or G3P) and anaerobic (dihydroxyacetone or DHA) pathways of catabolism. Enzyme and transport activities of the aerobic pathway are elevated in cells grown under oxygenated conditions on glycerol or G3P. Anaerobic growth on G3P as carbon source requires the presence of an exogenous hydrogen acceptor such as fumarate; cells thus grown also are highly induced in the G3P pathway. Anaerobic growth on glycerol requires no exogenous hydrogen acceptors; cells thus grown are highly induced in the DHA pathway but almost uninduced in the G3P pathway and the addition of fumarate electron acceptors has no effect on the relative levels of the two pathways. When both glycerol and G3P are provided anaerobically with fumarate, the DHA pathway is still preferentially induced, which probably accounts for the exclusive utilization of glycerol until its exhaustion. These observations suggest the presence of a regulatory control of G3P pathway imposed by the operation of the DHA pathway.
Effect of Temperature on Histidine Ammonia-Lyase from a Psychrophile,
Pseudomonas putida
Authors: Hug Daniel H. , Hunter John K.
Published: January 2020

Pseudomonas putida
was able to grow at 0 C in a complex medium containing
l
-histidine and to synthesize histidine ammonia-lyase and urocanase. The activity of the former enzyme was assessed between −10 and 60 C in cells and in cell extracts. Activity was maximal from 20 to 35 C. Below 20 C, activity decreased with temperature but, significantly, the enzyme exhibited 30% of its maximal activity at 1.5 C. The temperature response was similar in both intact cells and cell extracts, which indicated that the cell membrane did not significantly limit the entry of histidine at low temperature. Above and below the maximal temperature range, the reduced activity was not caused by irreversible inactivation, as shown by preincubation experiments. Also, when the temperature was rapidly changed from 60 to 30 C during an assay, the reaction rate increased abruptly to the full 30 C activity without a lag. This demonstrated the rapid reversibility of inactivation. The apparent Michaelis constant increased with temperature. As the substrate concentration was decreased, the enzyme activity became less dependent on temperature. The efficiency of substrate entry and catalysis near 0 C are factors in the ability of this facultative psychrophile to grow in a histidine medium at 0 C.
Streptomycin-Induced Synthesis of Abnormal Protein in an
Escherichia coli
Mutant
Authors: Pinkett M. Olivia , Brownstein Barbara L.
Published: January 2020

To determine directly the effects of streptomycin on translational fidelity in intact cells, we studied the synthesis of β-galactosidase and of the coat protein of bacteriophage R17 in an
Escherichia coli
mutant in which the bactericidal effects of streptomycin are delayed. After the addition of streptomycin to exponentially growing mutant cells, protein synthesis continues at an undiminished rate for approximately an hour; however, as measured by enzyme assays, little functional protein is produced. Serological assays designed to detect β-galactosidase and bacteriophage R17 coat protein show that substantial amounts of the protein synthesized can react with antisera prepared against active β-galactosidase and phage R17, indicating the aberrance of the protein produced in the presence of the antibiotic. The polypeptides synthesized in the presence of streptomycin are degraded in the cell to a much greater extent than protein synthesized in the absence of the antibiotic. The proteolytic attack on this protein is not affected by inhibitors of serine proteases, suggesting that enzymes other than those involved in “normal turnover” of cellular protein are responsible. In this strain, certain of the multiple effects of streptomycin are separated in time and the production of abnormal protein (enzymatically inactive and susceptible to proteolytic attack) could be studied in the absence of the lethal effect of the drug.
Isolation of Mutants Defective in α-Amylase from
Bacillus subtilis
: Genetic Analyses
Authors: Yamaguchi Kazuo , Nagata Yoshiho , Maruo Bunji
Published: January 2020

The rate of α-amylase (EC 3.2.1.1) synthesis in
Bacillus subtilis
is regulated by a gene,
amyR
, located near a structural gene,
amyE
, for the enzyme. To construct a fine map of the
amyR-amyE
region, we isolated 28 mutants defective in α-amylase activity. Eleven mutants out of 28 showed no α-amylase activity, whereas the other 17 showed less α-amylase activity than the parent. Out of 17 partially positive α-amylase mutants, 10 produced temperature-sensitive enzymes, and 4 produced immunologically altered enzymes, two of which are concurrently temperature-sensitive, and 5 produced smaller amounts of α-amylases which are indistinguishable from normal enzyme in their temperature sensitivity and immunological properties. Two out of 11 α-amylase-negative mutants produced material that cross-reacted with anti-amylase serum, and 3 mutants carried suppressible mutations by the suppressor described by Okubo. Mapping data indicate that all 28 mutation sites are located in the
amyE
region, and none of the groups of the mutants mentioned above contains lesions that are clustered in a single region of
amyE
. The
amyR
gene seems most likely to adjoin the terminal region of
amyE
.
Bacterial Cell Division Regulation: Characterization of the
dnaH
Locus of
Escherichia coli
Authors: Filip Camille C. , Allen Jane Smith , Gustafson Ralph A. , Allen Robert G. , Walker James R.
Published: January 2020

The
dnaH
locus is the fourth gene to be identified as required for deoxyribonucleic acid polymerization in
Escherichia coli
. A temperature-sensitive mutant defective in this gene exhibited an abrupt decrease in rate of deoxyribonucleic acid synthesis when shifted to 42 C. The locus mapped in the
proC-purE
region of the chromosome by conjugation and was co-transducible with
purE. dnaH
+
is carried on the F′
13
episome and is dominant over the
dnaH
−
mutation.
Initiation of Spore Germination in
Bacillus subtilis
: Relationship to Inhibition of
l
-Alanine Metabolism
Authors: Prasad Chandan
Published: January 2020

The inhibitory effects of anthranilic acid esters (methyl anthranilate and
N
-methyl anthranilate) on the
l
-alanine-induced initiation of spore germination was examined in
Bacillus subtilis
168. Methyl anthranilate irreversibly inhibited alanine initiation by a competitive mechanism. In its presence, the inhibition could be reversed only by the combined addition of
d
-glucose,
d
-fructose, and K
+
. Both
l
-alanine dehydrogenase and
l
-glutamate-pyruvate transaminase, enzymes which catalyze the first reaction in
l
-alanine metabolism, were competitively inhibited by methyl anthranilate. The

K
i

values for germination initiation (0.053 mM) and of
l
-glutamate-pyruvate transaminase (0.068 mM) were similar, whereas that for
l
-alanine dehydrogenase (0.4 mM) was six to seven times higher. Since a mutant lacking
l
-alanine dehydrogenase activity germinated normally in
l
-alanine alone, it is speculated that the major pathway of
l
-alanine metabolism during initiation may be via transmination reaction.
Cross-Pathway Regulation: Histidine-Mediated Control of Histidine, Tryptophan, and Arginine Biosynthetic Enzymes in
Neurospora crassa
Authors: Carsiotis M. , Jones Richard F. , Wesseling Anne C.
Published: January 2020

In
Neurospora crassa
, histidine starvation of histidine mutants resulted in derepression of histidine, tryptophan, and arginine biosynthetic enzymes. The same tripartite derepression occurred in wild-type strain 74A when it was grown in medium supplemented with 3-amino-1,2,4-triazole, an inhibitor of histidine biosynthesis. Histidine-mediated derepression of tryptophan and arginine biosynthetic enzymes was not due to a lowered intracellular concentration of tryptophan or arginine, respectively. A discussion of possible mechanisms and of similar studies in prokaryotic and eukaryotic organisms is presented.
Biosynthesis of Peptidoglycan in
Gaffkya homari
: Role of the Peptide Subunit of Uridine Diphosphate-
N
-Acetylmuramyl-Pentapeptide
Authors: Hammes Walter P. , Neuhaus Francis C.
Published: January 2020

The incorporation of
N
-acetylmuramyl (MurNAc)-peptides from nucleotide-activated precursors (reference: uridine diphosphate [UDP]MurNAc-Ala
1
-
d
Glu
2
-Lys
3
-
d
Ala
4
-
d
Ala
5
) with incomplete or modified peptide subunits into peptidoglycan was studied with membrane preparations from
Gaffkya homari
. The effectiveness of their utilization at low and high concentrations was compared on the basis of the values of
V
max
/

K
m

and
V
max
, respectively. At low concentration, replacement of alanine by glycine in position 5 has a small effect on the activity of the peptidoglycan synthesizing system, whereas it has a significantly larger effect in positions 1 and 4. The importance of
d
-alanine in position 4 at low substrate concentrations is also observed with the incomplete UDP-MurNAc-peptides. For UDP-MurNAc-tripeptide and -tetrapeptide,
V
max
/

K
m

is 0.06 and 0.55, respectively, of the value for the -pentapeptide. At high substrate concentration, replacement of
d
-alanine by glycine in either position 1 or 5 decreases the activity to 0.37 of the value for the reference nucleotide, whereas replacement in position 4 has a smaller effect (0.74). The profiles established from
V
max
and
V
max
/

K
m

with UDP-MurNAc-tripeptide, -tetrapeptide, and -pentapeptide show good correlation. At low concentration the specificity profiles of phospho-MurNAc-pentapeptide translocase, catalyzing the initial membrane reaction, are similar to those for the peptidoglycan synthesizing system; at high concentration, however, the profiles differ. The translocase appears to provide a primary specificity barrier at high substrate concentration for UDP-MurNAc-Ala-
d
Glu-Lys-
d
Ala-
d
Ala and UDP-MurNAc-Ala-
d
Glu-Lys-Gly-
d
Ala, and at low concentration for UDP-MurNAc-Ala-
d
Glu-Lys and UDP-MurNAc-Ala-
d
Glu-Lys-Gly-
d
Ala. Moreover, it is suggested that an additional specificity barrier exists in the peptidoglycan synthesizing system for certain nucleotides. Thus, the cytoplasmic enzymes and the membrane-associated enzyme(s) cooperate to insure the formation of functioning peptidoglycan in this organism.
Properties of an Antigenic Polysaccharide from
Corynebacterium parvum
Authors: Dawes Joan , Tuach Sheena J. , McBride William H.
Published: January 2020

Corynebacterium parvum
strain 10390 is an antitumor agent and stimulant of the reticuloendothelial system and produces a soluble antigen towards the end of its growth cycle. This material, which is a cell wall component and can also be released from the organism by acid or alkaline hydrolysis, has been purified. It is an acidic polysaccharide of molecular weight 100,000 to 150,000 and contains galactose, glucose, fucose,
N
-acetylgalactosamine,
N
-acetylglucosamine, uronic acids, sialic acids, and a small proportion of amino acids. The antigen gives a precipitin reaction with antisera raised against the whole organism and also binds to animal cells. The antigenic determinants are extremely resistant to oxidation, reduction, and enzymatic and chemical hydrolysis, but the single cell-binding site is destroyed by alkali and also by
Helix pomatia
digestive juice, alginase, and neuraminidase without substantially affecting the molecular weight. This site is inaccessible until the molecule is released from the cell surface. The possibility that the soluble antigen is the biologically active fraction of
C. parvum
is discussed.
Observations on the Bisulfite Reductase (P582) Isolated from
Desulfotomaculum nigrificans
Authors: Akagi J. M. , Chan May , Adams Verna
Published: January 2020

The bisulfite reductase (P582) from
Desulfotomaculum nigrificans
was purified to homogeneity as judged by polyacrylamide gel electrophoresis. By colorimetric methods of analysis, the products of bisulfite reduction by this enzyme were determined to be trithionate, thiosulfate, and sulfide. Of these, trithionate was consistently found to be the major product, whereas the latter two were formed in lesser quantities. When [
35
S]bisulfite was incorporated as substrate, no labeled sulfide was detected. Furthermore, when trithionate and thiosulfate were isolated from reaction mixtures and chemically degraded,
35
S was found in all three sulfur atoms of trithionate; however, only the inner sulfur atom of thiosulfate was radioactive. From these data we conclude that the bisulfite reductase of
D. nigrificans
reduces bisulfite to trithionate and that thiosulfate and sulfide are endogenous side products of the reaction.
Isolation and Characterization of Rifampin-Resistant and Streptolydigin-Resistant Mutants of
Bacillus subtilis
with Altered Sporulation Properties
Authors: Sonenshein Abraham L. , Cami Brigitte , Brevet Jean , Cote Richard
Published: January 2020

Mutants of
Bacillus subtilis
with altered deoxyribonucleic-dependent ribonucleic acid polymerase activity have been isolated and characterized. These mutants, selected as strains resistant to rifampin or streptolydigin, demonstrate drug-resistant in vitro ribonucleic acid synthesis. Sporeforming ability and support of phage infection are altered in many of the mutants. Mutations to rifampin and streptolydigin resistance have been located on the
B. subtilis
chromosome and ordered relative to the markers
cysA14
and
str
.
Photoreactivation, Photoproduct Formation, and Deoxyribonucleic Acid State in Ultraviolet-Irradiated Sporulating Cultures of
Bacillus cereus
Authors: Baillie Elizabeth , Germaine G. R. , Murreli W. G. , Ohye D. F.
Published: January 2020

Photoreactivation of ultraviolet-irradiated
Bacillus cereus
T declined markedly during the development of stage IV forespores. During ultraviolet irradiation of a culture containing early and late stage IV forespores, both vegetative- and spore-type photoproducts were formed. The formation of vegetative-type photoproducts (mainly thymine dimers) decreased to nearly half during late stage IV, remaining constant until lysis of the mother cells began, when it fell to zero. Spore-type photoproducts were first observed during late stage IV and increased with the increase in numbers of late stage IV forespores. The occurrence of spore-type photoproducts preceded the development of refractile forespores by about 1 h. At stage III the nuclear material occupied a central position, and the ribosomes were at the periphery of the forespore protoplast. During stage IV the deoxyribonucleic acid (DNA) occurred in a peripheral position, and bundles of fibers (“transition” DNA) could be seen. By stage V, all of the DNA appeared to be of the spore type and was peripheral, and the forespore protoplast center was packed with ribosomes. Forespore stages II, III, and IV were classified by light and electron microscopy. The curve for electron microscope classifications preceded that for light microscope classifications by approximately one stage. The formation of spore-type photoproducts preceded differentiation of DNA by about 1 h, the latter coinciding with the development of refractility. Spore-type photoproducts have been associated with DNA in the A state, and the progressive change of the forespore DNA into this state is discussed in relation to the spore differentiation process.