bluuebunny/crossref_metadata_2025 huggingface dataset:
Inactivation of Herpes Simplex Virus by Concanavalin A
Authors: Ito Michio , Barron Almen L.
Published: January 2020

The infectivity of herpes simplex virus type 1 (HSV-1) was inactivated after treatment with either concanavalin A (ConA) or periodate. Phytohemagglutinin, wheat germ agglutinin, pokeweed mitogen, and neuraminidase failed to inactivate the virus. The effect of ConA could be specifically inhibited or reversed by the addition of α-methyl-
d
-glucoside or α-methyl-
d
-mannoside. Evidence was obtained that HSV-1 inactivated by ConA could adsorb to host cells. Viral aggregation was not a major mechanism in the inactivation of HSV-1 by ConA. Under the experimental conditions employed, inactivation of HSV-1 was faster by ConA than by antiserum and less temperature dependent. A ConA-resistant fraction was detected which appeared to adsorb less quickly than untreated virus, and penetration of ConA-resistant fraction was strikingly slow. The presence of aggregates in the virus preparation did not appear to account for the ConA-resistant fraction. Inactivation of viral infectivity by ConA was obtained only with enveloped viruses, since HSV-1, HSV-2, pseudorabies, and vesicular stomatitis virus were inactivated and vaccinia and echovirus type 6 were not.
Translation of
Pseudomonas aeruginosa
Bacteriophage PP7 RNA by a Cell-Free Amino Acid Incorporating System from
Escherichia coli
Authors: Bassel B. A. , Walsh E. C. , Chi Jia-Yen , Curry M. E. , Brown J. G.
Published: January 2020

We have compared the activities of the RNA genomes of
Pseudomonas aeruginosa
phage PP7 and coliphages Qβ and f2 in a cell-free amino acid incorporating system derived from
Escherichia coli.
The rate of incorporation of [
14
C]leucine in the PP7 RNA-directed system is greater than in the systems directed by either Qβ or f2 RNA. The response to changes in phage RNA concentrations is similar in all the systems, reaching a saturation level at 0.75 to 1.0 mg of RNA per ml of reaction mixture. Analysis of complete reaction mixtures of the PP7 RNA and of the Qβ RNA systems by sucrose gradient centrifugation shows generally similar patterns for both RNAs. The principal differences are that in the PP7 system a slightly higher percentage of RNA forms ribosome complexes and that the polysomes are somewhat smaller. PP7 RNA is also degraded more extensively during the reaction than is Qβ RNA. Analysis of the products of the reactions by acrylamide gel electrophoresis shows that PP7 coat protein is the only identifiable product of the PP7 RNA-directed system, suggesting that only the coat protein cistron is translated by
E. coli
ribosomes.
Coiled Rings of DNA Released from Cells Infected with Bacteriophages T7 or T4 or from Uninfected
Escherichia coli
Authors: Bernstein Carol , Bernstein Harris
Published: January 2020

The replicating intracellular DNA of phage T7 was labeled at high specific activity with tritiated thymidine. The DNA of uninfected
Escherichia coli
was similarly labeled. Portions of cells which contained replicating phage T7 or
E. coli
DNA were lysed by a lysozyme, freeze-thaw, sodium lauryl sulfate procedure, and the DNA was spread on Millipore membranes for visualization by autoradiography. The DNA of phage T7 appeared to be highly concatenated reaching lengths of up to 721 μm. Much of the DNA of phage T7 and
E. coli
was retained in compact globular structures. In addition, orderly coiled rings of varying diameter up to about 43 μm were regularly observed. Similar coiled ring structures were also observed in autoradiographs of replicating phage T4 DNA which had been prepared in previous experiments. Worcel and Burgi (27) have presented evidence that
E. coli
chromosomes, when gently extracted from cells, are in a multilooped and superhelically twisted configuration. The coiled rings which we have observed may correspond to the relaxed, multilooped configurations which they find when the superhelical twists have been relieved by one or more nicks in each loop.
Buoyant Density of the Hepatitis A Virus-Like Particle in Cesium Chloride
Authors: Feinstone S. M. , Kapikian A. Z. , Gerin J. L. , Purcell R. H.
Published: January 2020

We recently visualized by immune electron microscopy a virus-like particle in the stools of patients with hepatitis A. The particle measured approximately 27 nm in diameter and morphologically resembled a picornavirus or parvovirus. To further characterize this particle, we have determined its buoyant density in cesium chloride (CsCl) by ultracentrifugation. Hepatitis A particles from three positive stool specimens were isopycnically banded in separate experiments, and the gradient fractions were examined for particles by immune electron microscopy by using hepatitis A convalescent sera. In each experiment, the particles were observed in a normal distribution about a peak fraction with a mean density of approximately 1.4 g/cm
3
. The buoyant density of 1.4 g/cm
3
in CsCl together with its morphology and the reported resistance of hepatitis virus to acid, ether, and heat suggest that this particle is parvovirus-like.
Immunological Properties of Two Polypeptides of Mason-Pfizer Monkey Virus
Authors: Tronick Steven R. , Stephenson John R. , Aaronson Stuart A.
Published: January 2020

Two polypeptides were isolated from the Mason-Pfizer monkey virus (MP-MV). The polypeptides, designated MP-MV p26 and MP-MV p15, were 26,000 and 15,000 molecular weight, respectively, based on gel filtration chromatography in the presence of 6 M guanidine hydrochloride. Radioimmunoassays developed for MP-MV p26 and p15 were, respectively, 10 and 100 times more sensitive than immunodiffusion and 1 and 10 times more sensitive than micro-complement fixation for the detection of MP-MV. Antigens of other reverse transcriptase-containing RNA viruses did not cross-react in either MP-MV radioimmunoassay. Further, antisera against these other viruses did not react with the radioiodine-labeled MP-MV polypeptides. The MP-MV radioimmunoassays should be useful in studying the natural occurrence of this virus and its relationship to primate tumors.
Cleavage of Mengovirus Polyproteins In Vivo
Authors: Lucas-Lenard Jean
Published: January 2020

The synthesis of mengovirus-specific proteins in vivo was studied by labeling the viral proteins with radioactive amino acids under conditions in which host protein synthesis was almost completely inhibited. Pulse-chase experiments enabled the kinetic analysis of the cleavages of certain viral protein precursors and the formation of others. The pattern of cleavages of mengovirus precursor polypeptides is similar to that of encephalomyocarditis virus. The major difference between the two viruses seems to be in the molar concentration in which the various primary products are produced. The molar ratio of the A protein, which is the precursor of the capsid proteins, to that of the primary products F and C, is approximately 1.5 to 2.0: 1: 1. Possible explanations for the unequal appearance of the structural and nonstructural proteins are discussed.
Rhabdovirus Replication in Enucleated Host Cells
Authors: Wiktor T. J. , Koprowski H.
Published: January 2020

Infection of enucleated TC-7 monkey cells with rabies virus resulted in the synthesis of virus-directed RNA and the production of rabies antigens but not of infectious virus. The yield of infectious vesicular stomatitis virus from enucleated TC-7 cells, on the other hand, was almost as high as that from intact cells. Inhibition of the mitochondrial functions of enucleated cells by treatment with ethidium bromide did not influence the development of rabies antigens or the production of infectious vesicular stomatitis virus.
Growth in Agarose of Human Cells Infected with Cytomegalovirus
Authors: Lang David J. , Montagnier Luc , Latarjet Raymond
Published: January 2020

After infection by human cytomegalovirus (CMV), human diploid fibroblasts could grow in agarose medium for several generations. Clones of infected cells grew for weeks, although in every case they ultimately underwent lysis owing to the cytopathic effect of the virus. Virus was inoculated at high dilution and after UV irradiation in an effort to derive cells infected with noninfectious defective particles still capable of inducing cell stimulation. Dilute or irradiated virus occasionally yielded large colonies of replicating cells, although permanent transformation was not observed. One clone derived from UV-CMV-infected cells was passaged four times before undergoing lysis. During these passages the cells exhibited alterations in morphology and orientation.
Incomplete and Aberrant Particles of Actinophage MSP2 from Lysates of
Streptomyces venezuelae
Authors: Jones Lily A. , Hahn Marianne L. , Taylor Alton R.
Published: January 2020

Lysates of actinophage MSP2, propagated on
Streptomyces venezuelae
S13, contain at least 10
11
PFU/ml. During purification by centrifugation methods and by adsorption chromatography, a number of types of aberrant and incomplete phage particles were seen by electron microscopy. Infectious MSP2 had a buoyant density in CsCl of 1.52 g/cm
3
and an absorbance at 260 nm relative to that at 280 nm (
A
260
/
A
280
) of 1.53. Empty capsids banded at 1.276 g/cm
3
and partially filled capsids banded at 1.351 g/cm
3
, and
A
260
/
A
280
ratios were 0.77 and 1.24, respectively. Two kinds of light capsids found in CsCl fractions of 1.278 g/cm
3
probably include the 1.276 component. Some capsids were joined by tail-like structures. Ghosts and polyheads also were present. Aberrant particles observed by electron microscopy included two-tailed actinophage, phage with abnormal tail positions, and large-headed phage.
Properties of Feline Leukemia Virus II. In Vitro Labeling of the Polypeptides
Authors: Velicer Leland F. , Graves Donald C.
Published: January 2020

Radioactive labeling of proteins from guanidine-hydrochloride disrupted feline leukemia virus was done in vitro by attaching [
3
H]methyl groups to protein amino groups with reductive alkylation. When in vitro labeled disrupted virus was analyzed by gel filtration in the presence of 6 M guanidine-hydrochloride the expected six major and one minor protein peaks were detected. When the same preparation was analyzed by immunodiffusion, the three antigens tested were found to have retained their antigenic activity.
The DNA-Delay Mutants of Bacteriophage T4
Authors: Mufti Siraj , Bernstein Harris
Published: January 2020

Mutants of phage T4 defective in genes 39, 52, 58-61, and 60 (the DNA delay or DD genes) are characterized by a delay in phage DNA synthesis during infection of a nonpermissive
Escherichia coli
host. Amber (
am
) mutants defective in these genes yield burst sizes varying from 30 to 110 at 37 C in
E. coli
lacking an
am
suppressor. It was found that when DD
am
mutants are grown on a non-permissive host at 25 C, rather than at 37 C, phage yield is reduced on the average 61-fold. At 25 C incorporation of labeled thymidine into phage DNA is also reduced to 3 to 10% of wild-type levels. Mutants defective in the DD genes were found to promote increased recombination as well as increased base substitution and addition-deletion mutation. These observations indicate that the products of the DD genes are necessary for normal DNA synthesis. The multiplication of the DD
am
mutants on an Su
−
host at 37 C is about 50-fold inhibited if prior to infection the host cells were grown at 25 C. This suggests that a compensating host function allows multiplication of DD
am
mutants at 37 C in the Su
−
host, and that this function is active in cells grown at 37 C prior to infection, but is inactive when the prior growth is at 25 C. Further results are described which suggest that the products of genes 52, 60, and 39 as well as a host product interact with each other.
Replication of a Nuclear Polyhedrosis Virus in a Continuous Cell Culture of
Spodoptera frugiperda:
Purification, Assay of Infectivity, and Growth Characteristics of the Virus
Authors: Knudson D. L. , Tinsley T. W.
Published: January 2020

Nonoccluded virus, polyhedra, and occluded virus were purified from a continuous cell culture of
Spodopera frugiperda
infected with nuclear polyhedrosis virus. The optimal temperature for the replication and lateral transmission of infectivity for the nuclear polyhedrosis viruses (NPV) in cell culture was 27 C. End-point dilution and plaque assay procedures for the measurement of infectivity are described and compared. Dose-response data demonstrated that a single particle could initiate an infection, and the validity of the relationship of 0.7 PFU per mean tissue culture infective dose (TCID
5 0
) further substantiated the accuracy of these infectivity assays. Particle-infectious unit calculations gave a ratio of 62 to 310 nonoccluded virus particles TCID
5 0
. Growth cycle and lateral transmission experiments indicated that infectious material was released from cells 12 h postinfection (p.i.) and approached a maximal titer 4 days p.i. The number of polyhedra, nonoccluded virions, and TCID
5 0
produced per cell was also presented. Typical yields of NPV produced per liter flask suggested that insect cell culture systems represent a feasible means by which the replication of these viruses could be investigated.
Further Studies on the Replication of Rabies and Rabies-Like Viruses in Organized Cultures of Mammalian Neural Tissues
Authors: Matsumoto S. , Schneider L. G. , Kawai A. , Yonezawa T.
Published: January 2020

Organized cultures of mammalian spinal and dorsal root ganglions were used for a comparative study of the neurocytopathology caused by rabies and so-called rabies-like viruses. Electron microscopy and titration of infectivity revised earlier data in which no difference could be demonstrated between street and fixed-virus infection. In the present study, fixed virus produced inclusion bodies without apparent virus assembly. Sequential electron microscopy revealed that the main sites of virus assembly were the membranes of the Golgi complex. In contrast, rabies-like viruses freshly isolated from wild rodents produced inclusion bodies all of which were associated with virus replication. Electron microscopic evidence has led us to classify these strains as street virus. Nonneural cell elements from cultivated ganglions were susceptible to fixed virus and the cultures yielded higher titers of infectivity as compared to those of rabies-like viruses. Virus budding was shown to occur at the cell surface as well as at intracytoplasmic membranes.
Mechanism of Replication of φX174 Single-Stranded DNA IX. Requirement for the
Escherichia coli dnaG
Protein
Authors: McFadden Grant , Denhardt David T.
Published: January 2020

Escherichia coli
NY73, possessing a temperature-sensitive mutation in the
dnaG
locus, was rendered sensitive to bacteriophage φX174 by P1 transduction. φX174 reproduces in this strain at 30 C but not at 40 C. All three stages of φX174 replication, parental replicative form (RF) synthesis, RF replication, and progeny single-stranded DNA synthesis, are thermolabile in this mutant. Competition-annealing data show that both plus- and minus-strand synthesis are equally inhibited after shift up to 40 C during RF replication. We conclude that the
dnaG
gene product is required for the synthesis of both strands of φX RF during RF replication and of the complementary strand and viral progeny strands during stages I and III, respectively.
Proteolytic Cleavage of Subunits of the Nucleocapsid of the Paramyxovirus Simian Virus 5
Authors: Mountcastle Walter E. , Compans Richard W. , Lackland Henry , Choppin Purnell W.
Published: January 2020

The protein subunits of the nucleocapsid of the parainfluenza virus simian virus 5 isolated from infected cells after dispersion with trypsin, chymotrypsin, or ficin are cleaved proteolytically. The molecular weights of the subunits which result from cleavage depend on the enzyme used, but are around 43,000, compared to the native subunit of 61,000. In most instances cleavage of the subunit appears to be due to the protease used to disperse the cell, and follows cell disruption. Nucleocapsids composed of native, uncleaved subunits can frequently be obtained from infected cells dispersed without a proteolytic enzyme; however, cleavage occasionally occurs even under those conditions, indicating that cellular proteases can at times cleave this protein. Nucleocapsids containing uncleaved subunits can be isolated from cells persistently infected with simian virus 5, indicating that persistent infection is not invariably associated with intracellular cleavage of this protein. Nucleocapsids composed of native subunits are hydrophobic, whereas those composed of the cleaved subunit can be dispersed in aqueous solution. It is suggested that the portion of the molecule removed by cleavage may be responsible for a specific interaction during virus assembly between the nucleocapsid and those areas of plasma membrane which contain the non-glycosylated viral membrane protein, which is also hydrophobic. An amino acid analysis of native and cleaved subunits has been done. The portion of the subunit removed by cleavage does not have a high proportion of hydrophobic residues, suggesting that those present are arranged together to form a hydrophobic domain.

The N termini of both the native and cleaved subunits are blocked. This suggests that the portion of the molecule which is externally disposed and removed by cleavage contains the C terminus, and the cleaved subunit which reacts with the viral RNA contains the N terminus.
Temperate Bacteriophage Infectious for Asporogenic Variants of
Bacillus pumilus
Authors: Bramucci Michael G. , Lovett Paul S.
Published: January 2020

Bacillus pumilus
strain NRRL B-3275 is lysogenic for an inducible, nondefective temperate bacteriophage φ75. φ75 infects and lysogenizes several asporogenic mutants of
B. pumilus
strain NRS 576 but does not productively infect the spore
+
parent. φ75 DNA is a linear duplex with a mol wt of about 29 × 10
6
and a buoyant density of 1.701 g/cm
3
. The location of the φ75 prophage attachment site on the chromosome of both host strains is adjacent to a lysine marker. The apparent order is φ
75 att lys trp.
Murine Intracisternal A Type Particles Fail to Separate from the Membrane of the Endoplasmic Reticulum
Authors: Perk Kalman , Dahlberg John E.
Published: January 2020

Analysis of serial sections of murine cells containing intracisternal A particles revealed that over 99% of all A particles remain in a budding configuration. This indicates that these particles fail to detach from the membrane of the endoplasmic reticulum. This observation explains how, despite their intracellular abundance in certain murine tumors, no extracellular A-type particles can be found.
Molecular Weight of Two Oncornavirus Genomes: Derivation from Particle Molecular Weights and RNA Content
Authors: Bellamy A. R. , Gillies S. C. , Harvey J. D.
Published: January 2020

Sedimentation analysis and intensity fluctuation spectroscopy have been used in conjunction with the Svedberg equation to determine the particle molecular weights of Rous sarcoma virus (Prague strain) and avian myeloblastosis virus (BAI strain). The molecular weights of these two viruses are (294 ± 20) × 10
6
and (256 ± 18) × 10
6
, respectively. Values for the molecular weight of the RNA contained in each particle have been calculated as (5.58 ± 0.5) × 10
6
and (5.88 ± 0.5) × 10
6
. Since the proportion of the viral RNA represented by 4 to 7
S
low-molecular-weight material is known, the molecular weight of the 60 to 70
S
genomes may be calculated to lie in the range (3.8 ± 0.3 to 4.8 ± 0.4) × 10
6
for both particles. These estimates for the molecular weight of the 60 to 70
S
genome are much lower than previous estimates and fall within the range of current estimates of the size of a single 35
S
subunit. The implications of this finding are discussed in terms of current theories for the structure of the genome of RNA tumor viruses.
Levels of Rat Cellular RNA Homologous to Either Kirsten Sarcoma Virus or Rat Type-C Virus in Cell Lines Derived from Osborne-Mendel Rats
Authors: Scolnick Edward M. , Maryak Jean M. , Parks Wade P.
Published: January 2020

DNA transcripts from V-NRK and RT21c rat type-C viruses were found to differ in their sequence homology to Kirsten and Harvey sarcoma viruses. V-NRK DNA transcripts consistently had homology to Kirsten and Harvey sarcoma virus, whereas RT21c DNA transcripts did not. To explain the differences, the nucleic acids and structural proteins of the two type-C viruses, released from each of two cell lines derived from Osborne-Mendel rats, were analyzed by molecular hybridization and competition radioimmunoassays. The p30 and p12 structural proteins of the two viruses were found to be highly related immunologically. In the V-NRK virus preparation, two sets of distinct RNA sequences were found in approximately equal amounts. One set is homologous to Ki-SV, and the other homologous to RT21c. In contrast, the RT21c virus preparation was found to contain a different ratio of these sequences. In this case the RT21c-like RNA sequences are present in 100-fold excess as compared to the additional Ki-SV specific sequences. Both NRK and RT21c cells contain in their DNA the full complement of Ki-SV homologous sequences, but NRK cells express much higher levels of these Ki-SV sequences in their RNA. These additional sequences, not homologous to RT21c, which are detected in uninfected NRK cellular RNA or V-NRK rat virus, could also be detected in the 60-70S RNA from a Moloney mouse type-C virus released from the NRK cells infected with the Moloney type-C virus. The results suggest that type-C viruses released from NRK cells incorporate species of RNA present in NRK cells which are homologous to Kirsten and Harvey sarcoma viruses. Either these sequences are of cellular origin, or rat cells contain two endogenous viruses with completely distinct nucleic acid sequences.
Role of Genetic Recombination in DNA Replication of Bacteriophage Lambda II. Effect in DNA Replication by Gene Delta
Authors: Barta K. , Zissler J.
Published: January 2020

We have studied the effect of delta mutations in phage lambda on DNA synthesis as assayed by the accumulation of λ DNA in infected cells. We find that delta mutants appear to generate somewhat less DNA than λ
+
in a
rec
+
host, suggesting the wild-type delta gene may act in DNA replication. An additional clue to delta function arises if replication is measured in the gamma-negative situation where concatemer formation is abortive. In this situation, the wild-type delta gene has an “inhibitory” effect on replication. A similar inhibitory effect on replication due to delta is observed after infection of P
2
lysogens. We conclude from these studies that the delta gene may act with alpha, beta, and gamma genes, possibly in a process affecting DNA replication.
Induction of Prophage SPO2 in
Bacillus subtilis
: Prophage Excision in the Absence of Bacterial or Bacteriophage DNA Synthesis
Authors: Arwert Fré , Rutberg Lars
Published: January 2020

Bacillus subtilis
lysogenic for SPO2 wild type was induced under conditions preventing synthesis of both bacterial and phage DNA. The infectivity of phage DNA in transfection is strongly decreased under these conditions, whereas the activity of single phage genes as measured by marker rescue with superinfecting phage is unaffected. DNA from induced cells was sedimented in neutral sucrose gradients. After induction, phage DNA was detected at a position in the gradients, which was different from the bulk of the bacterial DNA, corresponding to linear double-stranded DNA of about 25 × 10
6
daltons. Similar results were obtained with bacteria lysogenic for a SPO2 prophage carrying a DNA-negative mutation. No separation of phage and bacterial DNA activity was detected when chloramphenicol was present during the induction period. These experiments show that prophage SPO2 can excise from the bacterial chromosome without previous replication.
Hepatitis B Core Antigen: Immunology and Electron Microscopy
Authors: Barker Lewellys F. , Almeida June D. , Hoofnagle Jay H. , Gerety Robert J. , Jackson Daniel R. , McGrath Philip P.
Published: January 2020

Two distinct viral antigens are associated with the hepatitis B virus: the hepatitis B surface antigen (HB
s
Ag, Australia antigen) and the hepatitis B core antigen (HB
c
Ag). HB
s
Ag, purified from the serum of asymptomatic human HB
s
Ag carriers, and HB
c
Ag, purified from the liver of a chimpanzee acutely infected with hepatitis B virus, were examined by serological and immune electron microscopic methods. Antisera raised against HB
s
Ag reacted with the outer, surface component of the Dane particle and with the 20-nm spherical and tubular particles present in HB
s
Ag-positive serum, but not with the internal component of the Dane particle or with purified HB
c
Ag particles. Antisera raised against purified HB
c
Ag particles reacted with the internal component of the Dane particle and with HB
c
Ag, but not with the surface of the Dane particle or with the 20-nm spherical and tubular particles associated with HB
s
Ag. Purified HB
c
Ag particles, 27 nm in diameter, demonstrated distinct subunits. The infectious form of hepatitis B virus appears to be represented by the 42-nm Dane particle composed of a 27-nm nucleocapsid core component (HB
c
Ag) surrounded by an antigenically and morphologically distinct lipoprotein surface component (HB
s
Ag).
In Vivo and In Vitro Transcription of SPP1 DNA by Host RNA Polymerase
Authors: Milanesi G. , Melgara F.
Published: January 2020

Most (if not all) SPP1 RNA can be synthesized in infected cells in the presence of chloramphenicol, or in vitro by
Bacillus subtilis
RNA polymerase.
In Vitro Radioisotopic Labeling of Proteins Associated with Purified Polyoma Virions
Authors: McMillen Janis , Consigli R. A.
Published: January 2020

An in vitro procedure using [
14
C]formaldehyde and sodium borohydride was used to label the proteins of purified polyoma virions. This method, which labeled the three capsid proteins and the four internal histones of polyoma virus, did not alter the biological and biophysical characteristics of the virus. In addition to the seven viral proteins, five additional radioactive peaks were resolved on polyacrylamide gels. Four of these peaks could be attributed to growing the virus in the presence of serum. The fifth peak appears to be a nonhistone host-contributed protein made before infection.
Comparative biophysical studies of hepatitis B antigen, subtypes adw and ayw
Authors: Chairez R , Hollinger F B , Brunschwig J P , Dreesman G R
Published: January 2020

Comparative biophysical and biochemical analyses were performed on purified preparations of hepatitis B antigen (HBs Ag) subtypes adw and ayw, including isoelectric pH evaluations, analysis of the different morphological forms, molecular weight determinations, and analysis of the polypeptides by polyacrylamide gel electrophoresis, Both HBs Ag-positive plasma and purified HBs Ag were analyzed by electrofocusing in a sucrose ampholyte gradient. Four distinct populations of HBs Ag with a pH range of 4.5 plus or minus 0.1 to 5.4 plus or minus 0.1 for unfractionated plasma samples and 3.9 plus or minus 0.05 to 4.9 plus or minus 0.05 for purified samples were detected in both adw and ayw preparations. Electron microscopic studies of each population of purified HBs Ag revealed 19- to 27-nm spheres in each fraction. Purified material labeled with 125I by the chloramine-T method behaved as one major population with an isoelectric pH value of 3.9 plus or minus 0.1. Purified adw preparations revealed a major population with a molecular weight of 3.7 times 10-6 and a second one of 4.6 times 10-6. Purified preparations of ayw contained one population with a molecular weight of 4.6 times 10-6. Polyacrylamide gel electrophoretic analysis of purified HBs Ag revealed nine polypeptides for ayw and seven for adw particles. These studies indicate that purified preparations of HBs Ag are heterogeneous and that distinct differences can be detected between the two subtypes.
Identification of the in vitro translation products of adenovirus mRNA by immunoprecipitation
Authors: Oberg B , Saborio J , Persson T , Everitt E , Philipson L
Published: January 2020

Adenovirus type 2 mRNA was translated in S30 extracts from Ehrlich ascites and wheat embryo cells. The in vitro products were identified by sodium dodecyl sulfate-gel electrophoresis after immunoprecipitation with specific antisera in the presence of urea. Seven virion polypeptides could be identified by immunoprecipitation. Three of these appear to be precursors to polypeptides of the virion. mRNA isolated late in adenovirus infection was separated into three size classes by zonal sedimentation. Material sedimenting at 26S was translated into polypeptides corresponding to the largest virion polypeptides II to IV, a 22S fraction corresponding to polypeptide V, and smaller polypeptides and a 15S fraction corresponding to polypeptide IX. A significant amount of polypeptide IX was also synthesized by the 26S and 22S RNA.
Vaccinia virus infection of HeLa cells. I. Synthesis of vaccinia DNA in host cell nuclei
Authors: LaColla P , Weissbach A
Published: January 2020

The replication of vaccinia virus is thought to take place exclusively in the cytoplasm of host cells. However, using DNA-DNA hybridization techniques, it can be shown that a significant fraction of the synthesis of vaccinia DNA takes place in the nucleus as well as the cytoplasm. The (3H) thymiding pulse-labeled vaccinia DNA synthesized in the nucleus reaches a maximum at about 3 h after infection, corresponding to the time of maximal DNA synthesis in infected cells. At this time host DNA synthesis drops to about 25% of the rate of the uninfected cells. Even with short labeling times (2 min) the nucleus is found to contain 60% of the incorporated (3H)thymidine, much of which is in vaccinia DNA. Prior inhibition of host nuclear DNA synthesis with mitomycin C, followed by removal of the antibiotic causes a subsequent inhibition of vaccinia DNA synthesis and complete suppression of mature virus. Purified nuclei, isolated from vaccinia-infected cells, also synthesize vaccinia DNA in vitro. Over 90% of the DNA synthesized in vitro by isolated nuclei contain vaccinia-specific sequences.
Structural proteins of simian virus 40. I. Histone characteristics of low-molecular-weight polypeptides
Authors: Pett D M , Estes M K , Pagano J S
Published: January 2020

The DNA-associated polypeptides of simian virus 40 (SV40), VP4 (mol wt 14,000), VP5 (mol wt 12,000), and VP6 (mol wt 11,000), have several properties characteristic of cell histones. After extraction from purified SV40 with dilute acids, these three polypeptides co-electrophoresed on low pH polyacrylamide gels with monkey-kidney cell histones F3, F2b, and F2a1. No virus polypeptide co-electrophoresed with histone F1. Polypeptides VP4, 5, and 6 lacked tryptophan, and only VP4 contained cysteine, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis of virus labeled in vivo with (3H)lysine and either (14C)tryptophan or (35S)cystine. All of the capsid polypeptides VP1, 2, and 3 contained tryptophan whereas only VP1 and 2 contained cysteine. In addition, VP4, 5, and 6 are rich in arginine and lysine when compared with virus labeled with a mixture of amino acids. Analysis of virus grown in cells labeled prior to infection showed that VP4, 5 and 6 were labeled fivefold greater than the major capsid polypeptide, VP1, which indicates that they were partially derived from preexisting cell histones. Based on these data and on previously determined molecular weight estimates, we conclude that VP4, 5, and 6 are histones F3, F2b, and F2a1, respectively, although the possibility that SV40 contains a small amount of F2a2 could not be excluded.
Ribonucleoproteins of Uukuniemi virus are circular
Authors: Pettersson R F , von Bonsdorff C H
Published: January 2020

The internal ribonucleoprotein (RNP) of Uukuniemi virus was released with Triton X-100 and analyzed on sucrose gradients. Three species of RNP sedimenting at 140 to 150, 105 to 120, and 85 to 90S could be separated. All of them contained the same ratio of core polypeptide (mol wt, 25,000) to RNA. Eelctron microscopy using rotatory shadowing showed that all three species were circular. Free ends were rarely seen. Measurements of the strands revealed three distinct length classes of about 2.8, 1.4 and 0.7 mu m. Polyacrylamide-agarose gels showed that the largest RNP contained the L RNA, the medium-sized RNP the M RNA, and the smallest RNP the S RNA.
pH Stability Studies with Avian Infectious Bronchitis Virus (Coronavirus) Strains
Authors: Cowen B. S. , Hitchner S. B.
Published: January 2020

A comparison of 17 infectious bronchitis virus strains, using the same test procedure and assay system, demonstrated that stability at an acid pH is a variable characteristic of the avian coronaviruses.
Electron microscopy of simian virus 40 DNA configuration under denaturation conditions
Authors: Mayer F , Mazaitis A J , Pühler A
Published: January 2020

After isolation, the DNA of simian virus 40 appeared as a negative supertwist (form I) or as an open circle with at least one single-strand scission (form II). Under the denaturation conditions usually applied, such as heating in the presence of formaldehyde or application of alkali, form I molecules could appear as "relaxed" circles without single-strand scissions (form I') containing denatured sites not visible under the electron microscope. Form II molecules, under these denaturation conditions, showed partial or complete strand separations allowing the construction of denaturation maps. By using a modified denaturation procedure, i.e., heating of isolated SV40 DNA in the presence of dimethyl sulfoxide and formaldehyde followed by keeping the DNA in this denaturation solution at room temperature for periods up to 3 weeks, partially denatured relaxed circles without single-strand scissions were produced (form I'D) in addition to completely denatured form II molecules. The absence of single-strand scissions in form I'D molecules was demonstrated by a second heat treatment, which did not change the configuration of this molecular form. Form I'D molecules, in contrast to form I', contained denatured sites clearly discerible under the electron microscope. This combined application of two subsequent denaturation steps (denaturation by heating followed by denaturation at room temperature and neutral pH) showed that the molecular configuration I'D originated in two steps. The heating procedure produced molecules not distinquishable by electron microscopy from form I. In contrast to form I, these molecules were assumed to possess "preformed" denaturation sites (form I). Further treatment of form I molecules with denaturation solution at room temperature finally transformed them into convalently closed, relaxed, partially denatured circles exhibiting strand separations easily measurable on electron micrographs (form I'D). Denaturation maps of form I'D molecules were constructed by computer and compared with denaturation maps derived from partially denatured form II molecules. From these denaturation maps it can be concluded that the melting of base pairs occurring during the transition of simian virus 40 DNA form I into form I'D also preferentially happened at sites rich in the bases adenosine and thymine.
Purification and properties of a bacteriophage T5-modified form of Escherichia coli RNA polymerase
Authors: Szabo C , Moyer R W
Published: January 2020

A modified form of Escherichia coli RNA polymerase that contains one of the three T5-specific polypeptides known to interact with the host enzyme was purified from bacteriophage T5-infected cells. The properties of this T5-modified enzyme appeared identical to those of the RNA polymerase derived from uninfected non-colicinogenic cells and to a fully active enzyme isolated from T5-infected ColIb+ cells after the limited in vivo transcription of T5 genes allowed by the plasmid had ceased.
In vitro polyoma DNA synthesis: inhibition by 1-beta-d-arabinofuranosyl CTP
Authors: Hunter T , Francke B
Published: January 2020

The effects of 1-beta-D-arabinofuranosyl CTP (ara-CTP) on DNA replication were studied in an in vitro system from polyoma-infected BALB/3T3 cells. Ara-CTP concentrations of larger than or equal to 150 muM were found to block in vitro DNA synthesis completely, and concentrations of smaller than or equal to 0.3 muM had no inhibitory effect. Intermediate concentrations resulted in a concentration-dependent reduction of the in vitro synthesis rate. Long-term labeling with [alpha-32-P]ara-CTP demonstrated the incorporation of the analogue into cellular and viral DNA concomitantly with [3-H]TTP. In pulse-labeling experiments, at noninhibitory concentrations of the analogue, ara-CTP was incorporated into short DNA fragments and long growing strands to relatively the same extent as TTP. Partial venom phosphodiesterase digestion liberated the incoporated are-CTP at essentially the same rate as incorporated TTP, excluding a predominantly terminal incorporation, and after total venom phosphodiesterase digestion greater than 80% of the incorporated ara-CTP was recovered as 5'-ara-CMP. Analysis of the long-term in vitro viral DNA product made in the presence of partially inhibiting ara-CTP concentrations demonstrated that none of the steps leading to mature viral DNA were totally inhibited at the ara-CTP concentrations used. Pulse labeling of replicating viral DNA in the presence of ara-CTP revealed two consistent differences in the pattern found in control pulses: (i) predominant labeling of short chains (5S) with reduced amounts of radioactivity in the longer growing viral DNA strands (smaller than or equal to 16S), and (ii) a one-third to one-half reduction in size for short DNA chains labeled in the presence of ara-CTP. Release of the ara-CTP inhibition with excess dCTP resulted in covalent extension of these smaller short chans to approximately the size of regular short chains labeled in the absence of the inhibitor. Isolated short chains synthesized in the presence of ara-CTP exhibited a slightly lower degree of self-complementarity than regular short chains. The predominant labeling of short chains during pulses is, therefore, not a consequence of discontinuous growth on both sides of the replication fork. Similar results were obtained with ara-ATP and N-ethylmaleimide. The experiments indicate that ara-CTP acts primarily on DNA-polymerizing activities, affecting different DNA polymerases to varying degrees. The results are discussed in terms of the possible number and identity of polymerases involved in viral (and cellular) DNA replication.
Purification and characterization of the DNA polymerase and RNase H activities in Moloney murine sarcoma-leukemia virus
Authors: Gerard G F , Grandgenett D P
Published: January 2020

Two RNase H (RNA-DNA hybrid ribonucleotidohydrolase, EC 3.1.4.34) activities separable by Sephadex G-100 gel filtration were identified in lysates of Moloney murine sarcoma-leukemia virus (MSV). The larger enzyme, which we have called RNase H-I, represented about 10% of the RNase H activity in the virion. RNase H-I (i) copurified with RNA-directed DNA polymerase from the virus, (ii) had a sedimentation coefficient of 4.4S (corresponds to an apparent mol wt of 70,000), (iii) required Mn-2+ (2 mM optimum) for activity with a [3-h]poly(A)-poly(dT) substrate, (iv) eluted from phosphocellulose at 0.2 M KC1, and (v) degraded [3-H]poly(A)-poly(dT) and [3-H]poly(C)-poly(dG) at approximately equal rates. The smaller enzyme, designated RNase H-II, which represented the majority of the RNase H activity in the virus preparation, was shown to be different since it (i) had no detectable, associated DNA polymerase activity, (ii) had a sedmimentation coefficient of 2.6S (corresponds to an apparent mol wt of 30,000), (iii) preferred Mg-2+ (10 to 15 mM optimum) over Mn-2+ (5 to 10 mM optimum) 2.5-fold for the degradation of [3-H]poly(A)-poly(dT), and (iv) degraded [3-H]poly(A)-poly(dT) 6 and 60 times faster than [3-H]poly(C)-poly(dG) in the presence of Mn-2+ and Mg-2+, respectively. Moloney MSV DNA polymerase (RNase H-I), purified by Sephadex G-100 gel filtration followed by phosphocellulose, poly(A)-oligo(dT)-cellulose, and DEAE-cellulose chromatography, transcribed heteropolymeric regions of avian myeloblastosis virus 70S RNA at a rate comparable to avian myeloblastosis virus DNA polymerase purified by the same procedure.
Transfection of Escherichia coli spheroplasts. V. Activity of recBC nuclease in rec+ and rec minus spheroplasts measured with different forms of bacteriophage DNA
Authors: Benzinger R , Enquist L W , Skalka A
Published: January 2020

The in vivo activity of the recBC nuclease was assayed by transfection of isogenic rec+ and rec minus spheroplasts with bacteriophage DNA of various origin and structure. The results indicate that the recBC nuclease can limit transfection at several stages during the production of an infective center; such limitations depend primarily on whether the DNA is in, or assumes, a nuclease-sensitive structure. The first stage of limitation can occur when a nuclease-sensitive transfecting molecule enters the spheroplast. Other potential limitation points occur during replication and maturation of the bacteriophage DNA. The initial stage can be bypassed by using recBC nuclease-resistant molecules such as circular forms. Through analysis of results with other DNA structures, we found that in vivo the effects of the double-strand exonucleolytic activity of the recBC nuclease predominated. The effects of the single-strand nuclease activities seem to be modified from those observed for the purified enzyme in vitro (Karu et al., 1974). Inside the cell, the single-strand exonuclease activity is very weak and the single-strand endonuclease activity is abolished almost completely.
Host-dependent restriction of mengovirus replication. II. Effect of host restriction on late viral RNA synthesis and viral maturation
Authors: Prather S O , Taylor M W
Published: January 2020

Restricted mengovirus replication in Mandin-Darby bovine kidney (MDBK) cells is characterized by a 400-fold reduction in infectious virus yield and a 40-fold increase in the production of noninfectious virus. Using conditions which insure that all MDBK cells are infected, virus-specific RNA and protein synthesis were measured in the restrictive host and in a permissive host for mengovirus, HeLa cells. Labeling kinetics and sucrose gradient analysis of mengovirus-specific RNA from MDBK cells show a reduction of 10-fold in virion RNA, 5-fold in double-stranded RNA, and 12.5-fold in single-stranded RNA. The viral RNA biosynthetic processes which occur late in the replicative cycle and result in the production of 90% of the single-stranded viral RNA that is packaged into capsid proteins in the permissive host are absent in restrictive MDBK cells. Viral protein synthesis as measured by labeled viral-specific polysome is decreased, and there is an accumulation of 80S subviral particles in the restricted host. It is suggested that restriction may act at a number of stages of viral replication and maturation.
Cellular adsorption function of the sialoglycoprotein of vesicular stomatitis virus and its neuraminic acid
Authors: Schloemer R H , Wagner R R
Published: January 2020

Exposure of vesicular stomatitis (VS) virions to neuraminidase resulted in loss of their ability to agglutinate goose erythrocytes and to attach to L cells concomitant with hydrolysis of sialic acid. These viral adsorptive functions were also destroyed by tryspsinization. Sialyl transferase resialylation in vitro of neuraminidase-treated VS virions restored their hamagglutinating and adsorptive functions almost to original levels. Erythrocyte and L cell receptors for attachment of VS virions were blocked by fully sialylated fetuin and by VS viral sialoglycopeptides. Smaller VS viral glycopeptides generated by extensive trypsinization were less effective inhibitors of hemagglutination than were larger glycopeptides; neuraminic acid and neuraminosyl lactose had no capacity to inhibit hamagglutination or adsorption of virus to L cells. These data suggest that cellular receptors for viral adsorption recognize sialoglycopeptides of a certain size. Neuraminidase desialylation did not significantly alter the isoelectric point of VS virions. Cells exposed to DEAE-dextran, trypsin, or neuraminidase showed significantly increased capacity to attach fully sialylated but not desialylated VS virions. Neuraminidase desialylation of L cells, Chinese hamster ovary cells, and Madin-Darby bovine kidney cells resulted in enhanced susceptibility to plaque formation by VS virus.
Characterization of new regulatory mutants of bacteriophage T4. II. New class of mutants
Authors: Chace K V , Hall D H
Published: January 2020

New mutants of bacteriophage T4 that overproduce the enzyme dihydrofolate reductase were investigated. Unlike previously described overproducers of this enzyme (Johnson and Hall, 1974), these mutants did not overproduce deoxycytidylate deaminase. Overproduction of dihydrofolate reductase by the new mutants occurred because enzymatic activity continued to increase for a longer period of time in cells infected by the mutants than in cells infected by wild-type phage. This continued increase occurred even in the presence of rifampin, indicating that the overproduction is probably due to a post-transcriptional event. Both these new overproducers and the previously described overproducers were studied by using polyacrylamide gel electrophoresis. The two types of overproducers appeared to be very different. The previously described overproducers showed a delay and/or reduction in the synthesis of several proteins that normally started to be made 4 to 6 min after infection. Several proteins could be seen to be overproduced on the gels. The new overproducers did not show the delay in the synthesis of some proteins and only overproduced a few proteins. The new gene defined by the new overproducers is between the gene coding for thymidine kinase and the gene coding for lysozyme.
Assay for Epstein-Barr virus based on stimulation of DNA synthesis in mixed leukocytes from human umbilical cord blood
Authors: Robinson J
Published: January 2020

Relationships between the rate of DNA synthesis in cultured human umbilical cord leukocytes and the multiplicity of added Epstein-Barr virus (EBV) were studied. At low multiplicities of approximately 0.1 transforming units/cell (approximately 10 physical particles/cell), inoculated cultures demonstrated increased rates of DNA synthesis, by comparison to uninoculated cultures, 3 days after inoculation. Stimulation of DNA synthesis was evident of progressively longer intervals after inoculations of 10-fold dilutions of virus. The rate of DNA synthesis, determined by short [-3H]thymidine pulses, reflected as small as twofold changes in multiplicity and thus can serve as a quantitative assay for the virus. Changes in the rate of DNA synthesis were evident before increases in cell number or alteration in morphology. Stimulation of DNA synthesis in umbilical cord leukocytes was inhibited by treatment of EBV with antibody and also in graded fashion, by progressive doses of UV irradiation to the virus. Induction of DNA synthesis by EBV was serum dependent. Estimates of the number of cells transformed were obtained by extrapolation from a standard curve relating known numbers of transformed cells to [-3H]thymidine incorporation and also by cloning cells after exposure to virus. At the low multiplicities of infection used in these experiments approximately 0.04 to 0.002 of the total cellular population was transformed. The high efficiency of cell transformation by EBV by comparison to other DNA tumor viruses is emphasized.