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R C Clowes

Publications and source records attributed to R C Clowes.

27 records · Page 2Linked to original sources
PubMed1971

Mitomycin C and temperature induction of colicin B in the absence of deoxyribonucleic acid synthesis.

An Escherichia coli K-12 strain and its mutant, temperature-sensitive in deoxyribonucleic acid (DNA) synthesis, were used as hosts for two different ColB factors (ColB3-K166 and ColB4-K98). Induction of either colicin occurred in both hosts in the presence of mitomycin C at 42 C. Induction of the temperature-sensitive colicinogenic hosts also occurred without mitomycin C at 42 C at which temperature no DNA synthesis was observed. Colicin synthesis was a nonlethal event in ColB4-K98(+) cultures, whereas in cultures of ColB3-K166(+) induction led to death of a large fraction of the population.

Colicins↗
PubMed1971

ColB2-K77, a fertility-repressed F-like factor.

The colicinogenic B factor, transferred from Escherichia coli strain K77 (and termed ColB2-K77 or ColB2) to an E. coli K12 F(-) strain, is capable of promoting its own transfer to other K12 F(-) strains at a low rate (from LFC cultures) which can be increased under special conditions (HFC cultures). LFC cultures of K12 (ColB2)(+) F(-) strains show a low level of adsorption of F-specific phage particles which also increases under HFC conditions. The ColB2 factor is thus inferred to be an F-like sex factor which is repressed in its fertility. This repression is concluded to be due to a cytoplasmic repressor since, when ColB2 is present in cells containing an F factor (either autonomous or integrated), F fertility is also repressed as shown by the inability of such (ColB2)(+)F(+) [or (ColB2)(+)Hfr] strains to plaque F-specific phages, and by a reduction in the level of chromosomal transfer from such strains, compared to the corresponding F(+) (or Hfr) control strains. Mutants of the ColB2 factor in which fertility is no longer repressed (fertility derepressed or Fdr mutants) have been isolated. The ColB2Fdr mutant strains do not appear to be able to mobilize chromosomal transfer, although they have acquired F-specific phage sensitivity demonstrable by plaque formation and they transfer their colicin factor at high frequency and are well piliated. The Fdr mutation is presumed to result in the inability to synthesize the cytoplasmic fertility repressor since the ColB2Fdr factor does not repress the fertility of an F factor when present in the same host strain. A fertility-repressed drug resistance factor of the R(f) type is not stable in the presence of a ColB2 factor in the same cell and is eliminated in about 10% of the cells per generation. In contrast, another factor characteristic of the R(i) type is fully compatible with ColB2. Under conditions artificially stabilizing (ColB2Fdr)(+) (Rf)(+) strains, the enhanced fertility of ColB2Fdr is not repressed by the presence of the R factor, nor does the presence of R(f) in the intermediate strain of an HFC (for ColB2) system inhibit the normal increase in ColB2 transmissibility. It is concluded that the repressors of R(f) and ColB2, although both active on F fertility, are different; this may indicate that at least two independently repressible cistrons are involved in the expression of fertility characteristics.

Adsorption↗
PubMed1970

Molecular recombination between R-factor deoxyribonucleic acid molecules in Escherichia coli host cells.

THREE PREVIOUSLY STUDIED R FACTORS WERE USED: 222/R4, controlling transmissible resistance to sulfonamide, streptomycin, chloromycetin, and tetracycline (SU(r) SM(r) CM(r) TC(r)); 222/R3, a derivative of 222/R4 (now termed 222/R3W) having lost TC(r); and R15, controlling infectious resistance to SU and SM only. Two types of derivative R factors were isolated from 222/R4 by serial subculture in Salmonella species. One derivative, termed 222/R1, lost resistance to SU, SM, and CM, and the other, termed 222/R3N, lost only TC(r). Each factor was transferred to a standard Escherichia coli K-12 host. Recombinant factors of 222/R4 phenotype were isolated by selection after mixed culture of E. coli (222/R1)(+) and (222/R3N)(+) strains. Density-gradient equilibrium centrifugation of lysates of E. coli R(+) hosts in the presence of ethidium bromide separated R-factor deoxyribonucleic acid (DNA) as a heavy satellite peak which was subjected to electron microscopy or analytical density gradient centrifugation. Each DNA comprised a unimolecular species of circular DNA. The contour of R15 measured 22.3 mum [equivalent to 46 x 10(6) atomic mass units (AMU)], and that of 222/R4 measured 33.6 mum (70 x 10(6) AMU). 222/R3W appeared to be a point mutant or small deletion of 222/R4 with an almost identical size, whereas 222/R3N had lost a DNA segment of about 3 mum, and measured 30.3 mum or 63 x 10(6) AMU. The 222/R1 factors also appeared to have arisen by loss of DNA from 222/R4, 222/R1A being 22.3 mum or 46 x 10(6) AMU, whereas all other 222/R1 factors appeared to be duplicates, measuring 25.6 mum or 53 x 10(6) AMU. The DNA from six recombinant factors of R4 phenotype was indistinguishable in size and configuration from the parental 222/R4. In most cases, the number of R-factor copies (present as covalently closed circular molecules) per copy of the E. coli chromosome was less than 2, ranging from 1.2 to 3.3.

Centrifugation, Density Gradient↗
PubMed1970

Circular R-factor molecules controlling penicillinase synthesis, replicating in Escherichia coli under either relaxed or stringent control.

Two infectious drug-resistance (R) factors, R28K and R6K, each conferring resistance to a number of penicillins by the synthesis of a penicillinase, were transferred to Proteus mirabilis PM1 and Escherichia coli RC85 host strains. Deoxyribonucleic acid (DNA) extracted from these strains was separated by density-gradient centrifugation and subjected to electron microscopy by use of a modification of the protein-monolayer diffusion technique. Analytical density-gradient centrifugation of the purified DNA from PM1 strains showed, in addition to the major peak at a density of 1.698 g/cm(3) characteristic of Proteus chromosomal DNA, a single satellite band at a density of 1.710 g/cm(3) [guanine plus cytosine (GC) base ratio 50%] for R28K and at 1.704 g/cm(3) (GC base ratio 45%) for R6K. Direct CsCl density-gradient centrifugation of crude lysates of the E. coli (R28K)(+) strain in the presence of ethidium bromide gave rise to a sedimentation profile with a single satellite peak containing covalently closed circular (CCC) DNA molecules with a mean contour length of 21.4 mum [44 x 10(6) atomic mass units (AMU)], although a minority was 13.6 mum in length. From the size of the major class, it was estimated that there were two to three copies of the R28K factor present as CCC molecules per chromosome at various phases of cellular growth. Similar studies of the E. coli (R6K)(+) lysates showed two satellite peaks; peak I contained mostly CCC molecules of contour length 12.8 mum (26 x 10(6) AMU), and peak II, intermediate to peak I and the chromosomal peak, contained CCC molecules of a similar size, together with about equal numbers of catenated molecules, mostly dimers consisting of two interlocked monomers of 12.8 mum. A smaller number (ca. 0.1%) of higher catenanes was also seen. The number of CCC copies of the R6K factor per chromosome present in peak I was calculated as 13 at logarithmic phase and 38 at stationary phase. In peak II, a constant ratio of about one catenated dimer per chromosome was found at all phases of growth. Penicillinase assays of cultures at different phases of growth showed a correlation between the estimated number of R-factor copies present as CCC molecules and the enzyme activity per cell for both R28K and R6K.

Ampicillin↗
PubMed1967

A comparative study of two F-like colicin factors, ColV2 and ColV3, in Escherichia coli K-12.

Of three colicin factors, each determining the synthesis of a colicin V in three wild-type Escherichia coli strains studied, two were shown to have sex-factor activities. In E. coli K-12, these activities resembled those of the F sex factor (including rapid and efficient self-transmission in exponentially growing cultures, adsorption of "male-specific" ribonucleic acid phage, production of "female phenocopies," elimination by acridine orange, and chromosomal transfer dependent upon recombination with host bacterium) and differed in this way from those of the colicin I sex factor (ColI). The two V factors, ColV2 and ColV3, differed in their efficiency of plating male-specific phage and in the pattern of transfer of chromosomal markers. Furthermore, although neither factor could stably coexist with F within the same cell, they showed markedly different exclusion effects. In general, ColV2 excluded F and ColV3 was excluded by F, irrespective of which sex factor was preestablished in the cell. An exception to this was the ability of ColV2 to stabilize in any one of a series of Hfr strains, giving rise to strains which in the majority of cases showed normal Hfr and colicinogenic properties.

Acridines↗