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D J Cummings

Publications and source records attributed to D J Cummings.

At least 91 records · Page 5Linked to original sources
PubMed1972

Isolation and partial characterization of macro- and micronuclei from Paramecium aurella.

A method was developed for the isolation of macro- and micronuclei from Paramecium aurelia. This method utilized ionic and nonionic detergents to rupture the intact cells, calcium ions and spermidine were employed to protect the nuclei, and the nuclei were purified by centrifugation. Macronuclei consisted of 22% DNA, 10% RNA, and 68% protein. Micronuclei were composed of 9% DNA, 11% RNA, and 80% protein. DNA from both macro- and micronuclei had a density of 1.687 g/cc in CsCl and 1.417 g/cc in Cs(2)SO(4). These values corresponded to G + C content of about 23%. The RNA of macronuclei was examined by gel electrophoresis, and two high molecular weight species were identified having molecular Weights of 1.3 x 10(6) and 2.8 x 10(6) daltons. Three syngens were studied, and in each case the conditions for isolation of the nuclei were the same and no differences were observed in the properties of the nuclei.

Animals↗
PubMed1972

A remeasurement of the molecular weights of T-even bacteriophage substructural proteins.

T-even bacteriophage substructural proteins were studied by using discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was found that tail fibers are composed of two major proteins of 155,000 and 120,000 daltons molecular weight and four minor proteins of 51,000, 38,000, 27,000, and 23,000 daltons. Tail tubes were composed of one predominant protein of 18,500 daltons and one minor protein of 35,000 daltons molecular weight. Tubular polyheads obtained from a T4D amber mutant and by treatment of T4B-infected cells with L-canavanine were also examined, and no significant differences were noted in the molecular weight of the P23 protein.

Canavanine↗
PubMed1971

Head proteins from T-even bacteriophages. II. Physical and chemical characterization.

Three classes of structural head proteins, having molecular weights of 43,000, 18,000, and 11,000 daltons, were isolated from the T-even bacteriophages and were characterized. Based on electrophoretic studies, the 43,000-dalton class contained one major protein and one (or two) minor components, the 18,000-dalton class contained two protein components, and the 11,000-dalton class contained one major component. The N-terminal residues for the 43,000- and the 11,000-dalton classes were alanine, and the N-terminal residues for the 18,000-dalton class were methionine and alanine. Of the three classes of proteins, the 18,000-dalton proteins were the most acidic, whereas the 11,000-dalton proteins were the most basic. The amino acid composition of the 11,000-dalton class revealed that methionine and cysteine were absent and lysine, histidine, and tryptophan content was higher in the 11,000-dalton class than in the other two classes of proteins. Estimates of the relative number of the three classes of structural proteins were made and indicated that there were between 1,600 and 2,000 subunits of the 43,000-dalton proteins, 100 to 200 of the 18,000-dalton proteins, and 1,000 to 1,500 of the 11,000-dalton proteins. Evidence was presented that the 43,000-dalton proteins and the 11,000-dalton proteins readily formed aggregates with themselves but not with each other. The significance of these interactions to the structure of the T-even phage head was discussed.

Alanine↗
PubMed1971

Electron microscopic evidence for linear insertion of bacteriophage MU-1 in lysogenic bacteria.

Temperate bacteriophage Mu-1 was used to generate a lysogenic derivative of the F'lac episome of Escherichia coli. Intact, covalently circular molecules of F'lac and lysogenic F'lac Mu(+) deoxyribonucleic acid (DNA) were isolated and examined by electron microscopy. The mean contour lengths of F'lac and F'lac Mu(+) molecules were 37.6 +/- 0.4 mum and 53.2 +/- 0.4 mum, respectively. The mean difference, 15.6 mum, is similar to the mean contour length of 12.9 +/- 0.1 mum obtained for linear DNA molecules released by osmotic shock from mature phage Mu-1 virions. These results provide direct physical evidence that phage Mu-1 integrates by linear insertion of its genome into the DNA of lysogenic host bacteria. Chemical and physical analyses of phage Mu-1 DNA indicate that it is similar to E. coli DNA in respect of gross base composition, buoyant density, and melting temperature.

Adenine↗
PubMed1970

Thymineless death in Escherichia coli: deoxyribonucleic acid replication and the immune state.

Thymineless death (TLD) and nalidixic acid (NA) inactivation were studied in multiple auxotrophic strains of Escherichia coli B and B/r. As expected, it was found that both E. coli B and B/r exhibited an "immune state," i.e., a fraction of the population survived inactivation to both TLD and NA. With glucose as a carbon source in minimal medium, 0.1 to 0.3% of strain B and 0.2 to 0.5% of strain B/r survived inactivation; with acetate as the carbon source, the surviving fractions were increased to 1 to 2% and 5 to 7%, respectively. These immune fractions could be increased in magnitude by preincubation in minimal media containing thymine. Systematic analysis of the particular supplements necessary for the immune state indicated that the absence of the required amino acids was essential for the maximal expression of immunity. However, immunity was not abolished in acetate medium even in the presence of the required supplements. Further studies on the replication of deoxyribonucleic acid (DNA) during preincubation indicated that the degree of immunity did not necessarily correlate with the completion of a round of DNA replication. This finding was supported by examining the immune state in synchronous populations. In both glucose and acetate medium, there was no significant change in the degree of immunity to inactivation within the cell cycles of E. coli B and B/r. We concluded that some other event, possibly inhibition of protein synthesis, was necessary in determining the degree of the immune state. DNA replication was investigated after TLD and NA inactivation, and, as expected, it was found that both events led to premature initiation of replication. The only differences observed in the effects of these two processes on DNA synthesis were the following. (i) NA-induced replication was less sensitive to chloramphenicol than was TLD. (ii) TLD-induced replication was unaffected by pretreatment of the cells with mitomycin C, but this pretreatment prevented the replication of DNA after NA treatment. It was suggested that the mechanism of action of NA could involve a monofunctional attack on the DNA.

Acetates↗
PubMed1970

Head proteins from T-even bacteriophage. I. Molecular weight characterization.

T-even bacteriophage capsid proteins were separated on 6% agarose columns by use of 6 m guanidine hydrochloride containing 5 mm dithiothreitol both to dissociate and to elute the proteins. The head capsids of T2H, T4B, T4B01, T4D, and T6r(+) contained at least three structural proteins with molecular weights of 40,000, 18,000, and 11,000 daltons, amounting to 76, 2, and 8%, respectively, of the total capsid protein. On the other hand, T2L head capsids contained only two structural proteins with molecular weights of 40,000 and 18,000 daltons (81 and 2.5%, respectively, of the total protein). A discussion of the possible role of these structural head proteins and a T-even phage head model suggesting a structural arrangement of the 40,000 dalton subunit are presented.

Amino Acids↗
PubMed1970

Isolation and characterization of two basic internal proteins from the T-even bacteriophages.

Two species of basic internal proteins were found in osmotic shock supernatant solutions of bacteriophages T4B, T4D, T2H, T2L, and T6. The major species of protein isolated had a molecular weight of approximately 21,000 daltons, whereas the minor protein molecular weight was near 9,500 daltons. The two protein species exhibited unique isoelectric points and amino acid compositions. The 21,000-dalton protein of T2L showed major electrophoretic and compositional differences from the other 21,000-dalton proteins isolated. Similarities between the 21,000-dalton proteins and phage lysozyme are discussed.

Amino Acids↗
PubMed1970

Characterization of T-even bacteriophage substructures. I. Tail fibers and tail tubes.

T-even bacteriophages were grown and purified in bulk quantities. The protein coats were disrupted into their component substructures by treatment with 67% dimethyl sulfoxide (DMSO). Tail fibers and tubes were purified on glycerol-CsCl-D(2)O gradients and examined with respect to sedimentation properties, subunit molecular weights, amino acid composition, isoelectric points, and morphology. It was found that intact tail fibers had a sedimentation coefficient of 12 to 13S and that dissociated fibers consisted of three classes of proteins having molecular weights of 150 K +/- 10, 42 K +/- 4, and 28 K +/- 3 daltons. A model was constructed in which the 150-K subunit folded back on itself twice to give a three-stranded rope. Each 150-K subunit then represented a half-fiber and it was proposed that the role of the 42- and 28-K subunits was to hold each half-fiber together as well as serve as a possible link with other substructures. Isoelectric point studies also indicated that there were three different proteins with pI values of 3.5, 5.7, and 8.0. Amino acid analyses indicated that fibers had a composition distinct from other phage substructures. In addition, a striking difference was noted in the content of tryptophan among the phages examined. T4B had three to five times more tryptophan than did T2L, T2H, T4D, and T6. Intact tail tubes had an S(20,w) of 31 to 38S and dissociated tubes consisted of three proteins of molecular weights 57 K +/- 5, 38 K +/- 4, and 25 K +/- 3 daltons. Based on degradation studies with DMSO, it was proposed that these three proteins were arranged in a helical array yielding the tube structure. Isoelectric point studies indicated that there were three major proteins in the tube whose pI values were 5.1, 5.7, and 8.5. No significant differences were observed in the amino acid content of tubes obtained from all the T-even bacteriophages.

Amino Acids↗
PubMed1970

Characterization of T-even bacteriophage substructures. II. Tail plates.

Tail plates obtained from T4D amber mutants were examined with respect to sedimentation behavior, subunit molecular weights, amino acid composition, isoelectric points, and morphology. Intact plates had an S(20,w) of 77S from pH 5 to 9. The only conformational change noted was that below pH 5 tail plates readily dimerized yielding vis-à-vis dimers with an S(20,w) of 124S. Dissociated plates consisted of three major proteins with molecular weights of 53 K +/- 5, 31 K +/- 3, and 17 K +/- 2 daltons. The amino acid analyses indicated that plates had a composition distinct from fibers and tubes and were relatively rich in tryptophan. Degradation studies with dimethyl sulfoxide (DMSO) indicated that tail plates had a unique biological structure. After treatment with DMSO, and to some extent without DMSO, or from lysates of defective mutants, tetrad structures were observed in the electron microscope. These structures had an amino acid content and relative amounts of types of subunits similar but not identical to intact plates. It was proposed that plates were composed of nine such tetrads giving rise to a structure with six- and threefold symmetry.

Amino Acids↗
PubMed1969

Thymineless death in Escherichia coli: inactivation and recovery.

The effects of chloramphenicol (CAP) on the progress of thymineless death (TLD), nalidixic acid (NA) inactivation, ultraviolet (UV) irradiation, and mitomycin C (MC) inactivation were studied in Escherichia coli B, B(s-1), B(s-3), B(s-12), and B/r. This was done before, during, and after inactivation. During the progress of inactivation, it was found that at 10 to 20 mug of CAP per ml, up to 50% of the UV-sensitive bacteria survived TLD and about 10% survived NA. In E. coli B/r, at these concentrations of CAP, about 10 to 15% of the cells survived TLD and about 20 to 25% survived NA. Concentrations of CAP greater than 25 mug/ml actually increased the sensitivity of E. coli B, B(s-1), B(s-3), and B(s-12) to inactivation by either TLD or NA; at 150 mug of CAP per ml, the sensitivity of E. coli B/r to inactivation also increased. When E. coli B cells were incubated in CAP prior to inactivation, the longer the preincubation the longer onset of TLD was delayed; NA inactivation was also affected in that the rate of inactivation after CAP incubation was greatly decreased. Preincubation of E. coli B/r with CAP had much less effect on the progress of inactivation. After thymineless death, incubation in CAP plus thymine led to a rapid and almost complete recovery of E. coli B and B(s-12). Lesser recoveries were observed after inactivation due to UV, NA, or MC inactivation. E. coli B(s-1) and B/r did not recover viability after any mode of inactivation, and E. coli B(s-3) and B(s-12) recovered from UV to about 20% of the initial titer. It was suggested that protein synthesis, in particular proteins involved in deoxyribonucleic synthesis, was a determining factor in these inactivating and recovery events.

Bacterial Proteins↗
PubMed1968

Disruption of T-even bacteriophages by dimethyl sulfoxide.

Dimethyl sulfoxide (DMSO) disrupted T-even bacteriophages as well as lambda bacteriophage. The component substructures of T2L, T4B01, or T6, in particular heads, were readily isolated after treatment with 67% DMSO (v/v). In contrast, concentrations of DMSO above 50% not only separated heads from tails of bacteriophage lambda but led to degradation of the lambda heads. Examination of the isolated free heads of T-even bacteriophage indicated that a distinct neck substructure was attached to one apex of the head. On some free tails a similar neck substructure was also found at the proximal end of the sheath. The dimensions of this neck substructure were found to be about 130 by 180 A; by virtue of its size and morphological attachment to the free heads, it was concluded that this was a distinct substructure and not an extension of the tail tube.

Coliphages↗
PubMed1967

Thymineless death in Escherichia coli: strain specificity.

Thymineless death of various ultraviolet (UV)-sensitive strains of Escherichia coli B and K-12 was investigated. It was found that E. coli B, B(s-12), K-12 rec-21, and possibly K-12 Lon(-), all sensitive to UV, were also sensitive to thymine starvation. However, other UV-sensitive strains of E. coli were found to display the typical resistant-type kinetics of thymineless death. The correlation of these results with various other cellular processes suggested that the filament-forming ability of the bacteria might be involved in the mechanism of thymineless death. It was apparent from the present results that capacity for host-cell reactivation, recombination ability, thymine dimer excision, and probably induction of a defective prophage had little to do with determining sensitivity to thymine deprivation.

Coliphages↗