Herpes Simplex Virus: Genome Size and Redundancy Studied by Renaturation Kinetics

Herpes simplex virus subtype 1 deoxyribonucleic acid (DNA) was sheared in a French press to uniform fragments, denatured by heating, then allowed to reassociate. The renaturation reaction followed second-order kinetics with a single rate constant indicating that at least 95% of the genome was unique and that repetitive sequences, if present, were not detectable by this technique. The kinetic complexity of the herpes simplex genome was determined by DNA renaturation kinetics to be (95 ± 1) × 10⁶ daltons. Since this value is in excellent agreement with the molecular weight of viral DNA [(99 ± 5) × 10⁶ daltons] obtained from velocity sedimentation studies, it is concluded that virions contain only one species of double-stranded DNA molecules 95 × 10⁶ to 99 × 10⁶ daltons in molecular weight.     

The Enzymatic Synthesis of Rubber Polymer in Parthenium argentatum Gray

Washed rubber particles isolated from stem homogenates of Parthenium argentatum Gray by ultracentrifugation and gel filtration on columns of LKB Ultrogel AcA34 contain rubber transferase which catalyzes the polymerization of isopentenyl pyrophosphate into rubber polymer. The polymerization reaction requires Mg²⁺ isopentenyl pyrophosphate, and an allylic pyrophosphate. The K_m values for Mg²⁺, isopentenyl pyrophosphate, and dimethylallyl pyrophosphate were 5.2 × 10⁻⁴ molar, 8.3 × 10⁻⁵ molar, and 9.6 × 10⁻⁵ molar, respectively. The molecular characteristics of the rubber polymer synthesized from [¹⁴C]isopentenyl pyrophosphate were examined by gel permeation chromatography on three linear columns of 1 × 10⁶ to 500 Ångstroms Ultrastyragel in a Waters 150C Gel Permeation Chromatograph. The peak molecular weight of the radioactive polymer increased from 70,000 in 15 minutes to 750,000 in 3 hours. The weight average molecular weight of the polymer synthesized over a 3 hour period was 1.17 × 10⁶ compared to 1.49 × 10⁶ for the natural rubber polymer extracted from the rubber particles. Over 90% of the in vitro formation of the rubber polymer was de novo from dimethylallyl pyrophosphate and isopentenyl pyrophosphate. Treatment of the washed rubber particles with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate solubilized the rubber transferase. The solubilized enzyme(s) catalyzed the polymerization of isopentenyl pyrophosphate into rubber polymer with a peak molecular weight of 1 × 10⁵ after 3 hours of incubation with Mg²⁺ and dimethylallyl pyrophosphate. The data support the conclusion that the soluble preparation of rubber transferase is capable of catalyzing the formation of a high molecular weight rubber polymer from an allylic pyrophosphate initiator and isopentenyl pyrophosphate monomer.     

Biosynthesis of Novel Exopolymers by Aureobasidium pullulans

Aureobasidium pullulans ATCC 42023 was cultured under aerobic conditions with glucose, mannose, and glucose analogs as energy sources. The exopolymer extracts produced under these conditions were composed of glucose and mannose. The molar ratio of glucose to mannose in the exopolymer extract and the molecular weight of the exopolymer varied depending on the energy source and culture time. The glucose content of exopolymer extracts formed with glucose and mannose as the carbon sources was between 91 and 87%. The molecular weight decreased from 3.5 × 10⁶ to 2.12 × 10⁶ to 0.85 × 10⁶ to 0.77 × 10⁶ with culture time. As the culture time increased, the glucose content of the exopolymer extract formed with glucosamine decreased from 55 ± 3 to 29 ± 2 mol%, and the molecular weight increased from 2.73 × 10⁶ to 4.86 × 10⁶. There was no evidence that glucosamine was directly incorporated into exopolymers. The molar ratios of glucose to mannose in exopolymer extracts ranged from 87 ± 3:13 ± 3 to 28 ± 2:72 ± 2 and were affected by the energy source added. On the basis of the results of an enzyme hydrolysis analysis of the exopolymer extracts and the compositional changes observed, mannose (a repeating unit) was substituted for glucose, which gave rise to a new family of exopolymer analogs.     

Molecular Weight Determination of Sendai RNA by Dimethyl Sulfoxide Gradient Sedimentation

The molecular weight of the large RNA of Sendai virus has been determined by sedimentation analysis in sucrose gradients containing 99% dimethyl sulfoxide (DMSO) to be 2.3 × 10⁶. Sendai RNA recovered from 99% DMSO was found to cosediment with nondenatured Sendai RNA at 46 to 48s in ordinary sucrose gradients. The molecular weight value of 2.3 × 10⁶ is considerably smaller than the estimates of 6 × 10⁶ to 7 × 10⁶ determined under nondenaturing conditions, suggesting a unique structure for Sendai RNA.     

Physiochemical properties of the Vi antigen before and after mild alkaline hydrolysis

Sedimentation and diffusion constants were determined for electrophoretically purified Vi antigen before and after mild alkaline hydrolysis. The molecular weight of the intact antigen was found to be 1.7 × 10⁶. Mild alkaline hydrolysis completely removed O-acetyl, reduced the molecular weight to 1.0 × 10⁵, and lowered the intrinsic viscosity from 8.0 deciliters/g to approximately 0.5 deciliter/g. Infrared spectra of the two antigen preparations were also compared.     

Electron Microscopy of Viral RNA: Molecular Weight Determination of Bacterial and Animal Virus RNAs

A method for preparation of single-stranded RNA for electron microscopy determination of molecular weight is reported. The method uses treatment with formaldehyde at elevated temperatures to remove secondary structure and spreading in a protein monolayer from 50% formamide onto a 50% formamide hypophase. Molecular weights were determined for some bacterial and animal viruses, for which conflicting values had been reported earlier. Molecular weights determined by the method, using Escherichia coli large subunit rRNA for a standard (1.1 × 10⁶), are as follows: E. coli small subunit rRNA, 0.53 × 10⁶; coliphage f2-RNA, 1.3 × 10⁶; Qβ-RNA, 1.55 × 10⁶; and Newcastle disease virus RNA, 5.78 × 10⁶.     

Purification and Characterization of a Secreted Laccase of Gaeumannomyces graminis var. tritici

We purified a secreted fungal laccase from filtrates of Gaeumannomyces graminis var. tritici cultures induced with copper and xylidine. The active protein had an apparent molecular mass of 190 kDa and yielded subunits with molecular masses of 60 kDa when denatured and deglycosylated. This laccase had a pI of 5.6 and an optimal pH of 4.5 with 2,6-dimethoxyphenol as its substrate. Like other, previously purified laccases, this one contained several copper atoms in each subunit, as determined by inductively coupled plasma spectroscopy. The active enzyme catalyzed the oxidation of 2,6-dimethoxyphenol (K_m = 2.6 × 10⁻⁵ ± 7 × 10⁻⁶ M), catechol (K_m = 2.5 × 10⁻⁴ ± 1 × 10⁻⁵ M), pyrogallol (K_m = 3.1 × 10⁻⁴ ± 4 × 10⁻⁵ M), and guaiacol (K_m = 5.1 × 10⁻⁴ ± 2 × 10⁻⁵ M). In addition, the laccase catalyzed the polymerization of 1,8-dihydroxynaphthalene, a natural fungal melanin precursor, into a high-molecular-weight melanin and catalyzed the oxidation, or decolorization, of the dye poly B-411, a lignin-like polymer. These findings indicate that this laccase may be involved in melanin polymerization in this phytopathogen’s hyphae and/or in lignin depolymerization in its infected plant host.     

Comparison of the ribonucleic Acid subunits of reovirus, cytoplasmic polyhedrosis virus, and wound tumor virus

Double-stranded ribonucleic acid (RNA) from intact cytoplasmic polynedrosis virus (CPV) and wound tumor virus (WTV) was analyzed by polyacrylamide gel electrophoresis. Using RNA from type 3 reovirus as a standard, it was calculated that CPV-RNA consisted of 9 subunits corresponding to a molecular weight of 12.7 × 10⁶ and WTV-RNA consisted of 12 subunits corresponding to a molecular weight of 15.5 × 10⁶.     

Concentrations,Source and Risk Assessment of Polycyclic Aromatic Hydrocarbons in Soils from Midway Atoll,North Pacific Ocean

This study was designed to determine concentrations of polycyclic aromatic hydrocarbons (PAHs) in soil samples collected from Midway Atoll and evaluate their potential risks to human health. The total concentrations of 16 PAHs ranged from 3.55 to 3200 µg kg⁻¹ with a mean concentration of 198 µg kg⁻¹. Higher molecular weight PAHs (4–6 ring PAHs) dominated the PAH profiles, accounting for 83.3% of total PAH mass. PAH diagnostic ratio analysis indicated that primary sources of PAHs in Midway Atoll could be combustion. The benzo[a]pyrene equivalent concentration (BaP_eq) in most of the study area (86.5%) was less than 40 µg kg⁻¹ BaP_eq and total incremental lifetime cancer risks of PAHs ranged from 1.00×10⁻¹⁰ to 9.20×10⁻⁶ with a median value of 1.24×10⁻⁷, indicating a minor carcinogenic risk of PAHs in Midway Atoll.     

Inactivation of rubella virus by gamma radiation

The Gilchrist and M-33 strains of rubella virus exposed in the frozen state to ¹³⁷Ce or ⁶⁰Co were inactivated exponentially according to “one hit” kinetics. There was no difference in the radiosensitivity of the two strains. Experimental D₃₇ values for both strains ranged from 1.9 × 10⁵ to 2.9 × 10⁵ rads, and computed radiosensitive molecular weights ranged from 2.6 × 10⁶ to 4.0 × 10⁶ daltons.     

Plasmid Distribution and Evidence for a Proteinase Plasmid in Streptococcus lactis C2

Five plasmids, distinguishable by their molecular weights (10⁶, 2 × 10⁶, 5 × 10⁶, 10⁷, 3 × 10⁷, respectively) were isolated from Streptococcus lactis C2. A spontaneous proteinase-negative derivative of this strain lacked the 10⁷ plasmid.     

Bacterial Communities in Acidic and Circumneutral Streams

The relationship between pH and the abundance and activity of bacteria in streams was examined as part of a study of the effect of acidification on stream communities. Of the bacterial communities examined, the epilithic community appeared to be the most significantly affected by acidification. Microbial biomass, as quantified by measuring the ATP level, on rock surfaces was significantly correlated with pH. Also, bacterial production by the epilithic bacteria, indicated by incorporation of tritiated thymidine into DNA, was always higher at high-pH sites than at low-pH sites of the same stream order and elevation. Bacterioplankton concentrations varied between 0.53 × 10⁵ and 9.42 × 10⁵ cells · ml⁻¹ in the first- to fourth-order streams examined. The bacterioplankton concentration in one sample from a spring was 0.17 × 10⁵ cells · ml⁻¹. Bacterioplankton concentrations were not correlated with pH but were significantly correlated with seston concentrations. The correlation with seston is a result of increases in particle-associated bacteria at high seston concentrations. The proportion of bacterioplankton attached to particles varied from 0 to 70%. Bacterial numbers and production in the sediments were significantly correlated with the organic content of the sediment rather than with the pH of the overlying water. Thus, reduced abundance and activity of bacteria as a result of acidification could be detected only for the relatively active community on rock surfaces; this community was exposed to the low pH because of the unbuffered nature of its environment.     

Genome-Wide Association Study Identifies Major Loci for Carcass Weight on BTA14 in Hanwoo (Korean Cattle)

This genome-wide association study (GWAS) was conducted to identify major loci that are significantly associated with carcass weight, and their effects, in order to provide increased understanding of the genetic architecture of carcass weight in Hanwoo. This genome-wide association study identified one major chromosome region ranging from 23 Mb to 25 Mb on chromosome 14 as being associated with carcass weight in Hanwoo. Significant Bonferroni-corrected genome-wide associations (P<1.52×10⁻⁶) were detected for 6 Single Nucleotide Polymorphic (SNP) loci for carcass weight on chromosome 14. The most significant SNP was BTB-01280026 (P = 4.02×10⁻¹¹), located in the 25 Mb region on Bos taurus autosome 14 (BTA14). The other 5 significant SNPs were Hapmap27934-BTC-065223 (P = 4.04×10⁻¹¹) in 25.2 Mb, BTB-01143580 (P = 6.35×10⁻¹¹) in 24.3 Mb, Hapmap30932-BTC-011225 (P = 5.92×10⁻¹⁰) in 24.8 Mb, Hapmap27112-BTC-063342 (P = 5.18×10⁻⁹) in 25.4 Mb, and Hapmap24414-BTC-073009 (P = 7.38×10⁻⁸) in 25.4 Mb, all on BTA 14. One SNP (BTB-01143580; P = 6.35×10⁻¹¹) lies independently from the other 5 SNPs. The 5 SNPs that lie together showed a large Linkage disequilibrium (LD) block (block size of 553 kb) with LD coefficients ranging from 0.53 to 0.89 within the block. The most significant SNPs accounted for 6.73% to 10.55% of additive genetic variance, which is quite a large proportion of the total additive genetic variance. The most significant SNP (BTB-01280026; P = 4.02×10⁻¹¹) had 16.96 kg of allele substitution effect, and the second most significant SNP (Hapmap27934-BTC-065223; P = 4.04×10⁻¹¹) had 18.06 kg of effect on carcass weight, which correspond to 44% and 47%, respectively, of the phenotypic standard deviation for carcass weight in Hanwoo cattle. Our results demonstrated that carcass weight was affected by a major Quantitative Trait Locus (QTL) with a large effect and by many SNPs with small effects that are normally distributed.     

Sedimentation Coefficient and Fragility under Hydrodynamic Shear as Measures of Molecular Weight of the DNA of Phage T5

T5 DNA molecules resemble fragments of T2 DNA of molecular weight 84 × 10⁶ with respect to sedimentation coefficient and susceptibility to breakage under hydrodynamic shear. The sedimentation coefficient falls by the same factor when either T2 or T5 DNA is broken at its characteristic critical shear rate. At a given high rate of shear, both DNA's are broken into fragments exhibiting the same sedimentation coefficient. It follows that 84 × 10⁶ is a proper estimate of the molecular weight of T5 DNA, and that particles of phage T5, like those of T2, contain a single DNA molecule.     

Structure and Composition of Biological Slimes on Paper and Board Machines

Biological slimes (biofilms) collected from the wet end of paper and board machines were examined by electron microscopy and analyzed for fatty acid composition, neutral sugar composition, and ATP. Electron microscopy revealed minuscule prokaryotic organisms (diameter, 0.2 to 0.4 μm). Larger cells morphologically resembling Sphaerotilus and Leptothrix spp. were found in slimes from machines using recycled fiber or unbleached pulp. The bacteria were embedded in a slimy matrix and often contained reserve materials microscopically resembling poly-β-hydroxybutyrate and glycogen. Fatty acid analysis of the slimes revealed bacterial signature fatty acids in concentrations equivalent to the presence of 2 × 10¹⁰ to 2.6 × 10¹² (average, 7 × 10¹¹) bacterial cells (live and dead) per g (dry weight) of slime. The slimes contained several known components of bacterial polysaccharides in addition to glucose, indicating that the slime body consisted of bacterial polysaccharides. The slimes contained uronic acids equivalent to a binding capacity of 12.5 to 50 μmol of divalent cations per g (dry weight) of slime. The uronic acid-containing polysaccharides may be responsible for the accumulation of heavy metals in the slime. Calculation of the ATP contents of the slimes resulted in an estimate of 5 × 10¹² cells per g (dry weight) of slime when calibrated with pure bacterial cultures isolated from the slimes. From electron micrographs, an estimate ranging from 1 × 10¹⁰ to 1.5 × 10¹² (average, 4 × 10¹¹) cells per g (dry weight) of slime was obtained.     

The Molecular Weight and Other Biophysical Properties of Bromegrass Mosaic Virus

Data are presented which show that bromegrass mosaic virus has a particularly low molecular weight and nucleic acid content. A molecular weight of 4.6 × 10⁶ was calculated from the sedimentation coefficient, S°_20,w = 86.2S, the diffusion coefficient, D_20,w = 1.55 × 10^-7 cm²/sec., and an assumed partial specific volume, [UNK] = 0.708 ml/gm. The virus has a ribonucleic acid content of 1.0 × 10⁶ atomic mass units. Electrophoresis experiments showed that the virus is stable in 0.10 ionic strength buffers in the pH range 3-6. Breakdown of the virus was observed outside this pH range. Some characteristics of the breakdown products are described.     

Gene Expression During the Development of Bacillus subtilis Bacteriophage φ29 II. Resolution of Viral-Specific Ribonucleic Acid Molecules

The ribonucleic acid (RNA) specified by bacteriophage 29 was isolated under conditions which minimized physical and enzymatic degradation, reduced aggregation, and enriched for completed molecules. This RNA was fractionated both by sedimentation through sucrose density gradients and electrophoresis through polyacrylamide gels to measure the size and relative amount of each component. Early RNA consisted of six components of molecular weight 0.75 × 10⁶, 0.44 × 10⁶, 0.37 × 10⁶, 0.25 × 10⁶, 0.09 × 10⁶, and 0.04 × 10⁶, accounting for 35% of the coding capacity of 29 deoxyribonucleic acid (DNA). All of these components except the one at 0.44 × 10⁶ were detected when infection occurred in the presence of chloramphenicol. Synthesis of the major early component (0.75 × 10⁶) ceased shortly after the onset of viral DNA synthesis. The other species of early RNA were synthesized throughout the latent period. Three additional components, 1.75 × 10⁶, 0.93 × 10⁶, and 0.07 × 10⁶, appear at late times. The two large RNAs may be polycistronic messenger RNAs corresponding to the seven viral capsid proteins.     

Phanerochaete chrysosporium β-Glucosidases: Induction, Cellular Localization, and Physical Characterization

Phanerochaete chrysosporium produces intracellular soluble and particulate β-glucosidases and an extracellular β-glucosidase. The extracellular enzyme is induced by cellulose but repressed in the presence of glucose. The molecular weight of this enzyme is 90,000. The K_m for p-nitrophenyl-β-glucoside is 1.6 × 10⁻⁴ M; the K_i for glucose, a competitive inhibitor, is 5.0 × 10⁻⁴ M. The K_m for cellobiose is 5.3 × 10⁻⁴ M. The intracellular soluble enzyme is induced by cellobiose; this induction is prevented by cycloheximide. The presence of 300 mM glucose in the medium, however, had no effect on induction. The K_m for p-nitrophenyl-β-glucoside is 1.1 × 10⁻⁴ M. The molecular weight of this enzyme is ~410,000. Both enzymes have an optimal temperature of 45°C and an E_act of 9.15 kcal (ca. 3.83 × 10⁴ J). The pH optima, however, were ~7.0 and 5.5 for the intracellular and extracellular enzymes, respectively.     

Resolution and Properties of Two High Affinity Cyclic Adenosine 3':5'-monophosphate-Binding Proteins from Wheat Germ

A high affinity cAMP-binding protein (cABP II) was purified to homogeneity from wheat germ. The apparent molecular weight of cABP II, as determined from gel exclusion chromatography, is 5.2 × 10⁵ (at low ionic strength) and 2.8 × 10⁵ (at high ionic strength). One polypeptide subunit (molecular weight, 80,000) was resolved by polyacrylamide gel electrophoresis of cABP II under subunit dissociating conditions. The purification protocol employed resolves cABP II from a distinct, less acidic cAMP-binding protein (cABP I). The K_d values for cAMP are about 10⁻⁶ molar and 10⁻⁷ molar for cABP II and cABP I, respectively. The cAMP-binding sites of cABP I and cABP II have a marked adenine-analog specificity, binding adenine, adenosine, adenine-derived nucleosides and nucleotides and a variety of adenine derivatives having cytokinin activity. While cABP II is phosphorylated in reactions catalyzed by endogenous protein kinases, there is no evidence for modulation of these cABP II-protein kinase interactions by cAMP.     

Effects of Temperature and Pressure on Sulfate Reduction and Anaerobic Oxidation of Methane in Hydrothermal Sediments of Guaymas Basin

Rates of sulfate reduction (SR) and anaerobic oxidation of methane (AOM) in hydrothermal deep-sea sediments from Guaymas Basin were measured at temperatures of 5 to 200°C and pressures of 1 × 10⁵, 2.2 × 10⁷, and 4.5 × 10⁷ Pa. A maximum SR of several micromoles per cubic centimeter per day was found at between 60 and 95°C and 2.2 × 10⁷ and 4.5 × 10⁷ Pa. Maximal AOM was observed at 35 to 90°C but generally accounted for less than 5% of SR.