Search PubMed⌕ Search

Biomedical subjects

G Dougan

Publications and source records attributed to G Dougan.

263 records · Page 15Linked to original sources
PubMed1979

Structure and function of plasmid ColE1 and related plasmids.

Analysis of plasmid ColE1, its naturally occurring relatives ColK and CloDF13, and a wide range of ColE1 derivatives containing either insertions or deletions of genetic material has allowed localization on the ColE1 genome of DNA sequences responsible for colicin E1 synthesis, immunity to colicin killing, conjugal mobility and incompatibility. We have examined incompatibility between pairs of ColE1 derivatives ranging in size from 2.6 to 13.8 Md. Though all the plasmids tested exerted ColE1 incompatibility, a definite pattern was observed regarding the dominant plasmid in any pair tested (i.e. the plasmid that displaces the other from a heterozygote). Usually the larger plasmid is displaced. We conclude that loci for incompatibility reside within 0.7 kb of the ColE1 replication region. A model is presented to explain both the incompatibility data and the observation that the fraction of total DNA occurring as ColE1-like plasmid in a cell is approximately constant. Transposons Tn1 and Tn3 (3.2 Md; Apr and approximately 85% homologous), Tn501 (5.5 Md; Hgr), and Tn7 (9.3 Md; Tpr Smr) can all be transposed into ColE1. Though all have closely related. Tn501 and Tn7 do not complement transposition of Tn3 transposition defective deletions. A Tn3-specified 19,000 dalton protein is absent in one particular class of transposition-defective deletion.

Bacterial Proteins↗
PubMed1978

Mobilization of the Escherichia coli plasmid ColE1 (colicin E1) and ColE1 vectors used in recombinant DNA experiments.

The Escherichia coli Co1E1 plasmid, which codes for production of colicin E1, is inherently nontransferable (nonconjugative) by bacterial mating. Co1E1 can be transmitted at mating by a process called mobilization if Co1E1 is coresident with a transfer plasmid. Mobilization is governed in part by a Co1E1 gene called mob. Co1E1 is mob+. Several Co1E1 derivatives employed in recombinant DNA experiments, notably pBR313 and pBR322, are mob-. These cloning vehicles are mobilized at markedly reduced frequency relative to Co1E1. E. coli K12 carrying either pBR313 or pBR322 represents a useful host vector system for recombinant DNA experiments and affords a significant degree of biological containment.

Bacteriocin Plasmids↗
PubMed1978

Analysis of sequences transposed by complementation of two classes of transposition-deficient mutants of Tn3.

The Tn1 and Tn3 elements are closely related transposons which carry the structural gene for ampicillin resistance. Two classes of deletion mutants of the plasmid pMB8::Tn3 (RSF1050) are unable to transpose ampicillin resistance but can be complemented in trans by a coresident Tn1 or Tn3 element. The analysis of the sequences transposed upon complementation of one class of mutants (type I) showed that the mutant element had undergone bona fide transposition. Complementation of the type II mutants led to the transposition of a sequence analogous to bacteriophage mu-promoted integration of non-mu DNA. The transposed sequence consisted of two Tn3 elements which flanked a single copy of the pMB8 portion of the RSF1050 genome. Complementation data indicated that the type II mutants are defective in at least one trans-acting function which must be supplied for transposition to occur. The nature of sequence transposed from the type II mutant is the consequence of a defective cis-acting function (or site). In addition, the type II mutants were defective in a trans-acting function which regulated the frequency of transposition.

Base Sequence↗
PubMed1977

The transposon Tn1 as a probe for studying ColE1 structure and function.

Insertion of the transposable genetic element Tn1 into different sites of plasmid ColE1 results in a number of mutnat phenotypes. Whereas all plasmid examined were present in normal amount, all showed reduced immunity to killing by colicin E1. Of six insertions isolated after conjugation, five fail to produce colicin, are conjugally proficient (transmissible), and map within a 500 nucleotide region of the genome. The other is conjugally deficient, produces colicin normally and maps close to two others with a similar phenotype isolated after transformation. Of four others isolated after transformation, two have similar properties to the original five transmissible plasmids. The other two are nontransmissible and produce colicin. Non-transmissibility is correlated with reduced relaxation complex. Patterns of protein synthesis in minicells by ColE1 and ColE1 :: Tn1 plasmids have been examined: all ColE1 plasmids containing Tn1 show an altered pattern of ColE1 protein synthesis in addition to three presumptive Tn1-specified proteins, one of which is shown to be beta-lactamase. ColE1 :: Tn1 plasmids can be inserted into the conjugative plasmid R64drd11 to form a cointegrate in which ColE1 and Tn1 function can be expressed.

Colicins↗
PubMed1977

Changes in protein synthesis on mitomycin C induction of wild-type and mutant CloDF13 plasmids.

Mitomycin C treatment of Escherichia coli K-12 cells containing the nonconjugative plasmid CloDF13 resulted in inhibition of host chromosome protein synthesis and a high rate of synthesis of two CloDF13-specified proteins whose molecular weights correspond to cloacin and immunity protein. Five molecules of immunity protein were synthesized for each cloacin DF13 molecule. Mitomycin C-treated cells containing a copy mutant of CloDF13 made three to four times as much of each protein as cells containing wild-type CloDF13. CloDF13 plasmids that contained the transposon Tn1 were isolated. Two did not induce after mitomycin C treatment, failing both to inhibit host cell synthesis and to produce the two new proteins. In minicells, they showed reduced CloDF13-specified protein synthesis and produced three Tn1-specified proteins.

Bacteriocins↗
PubMedDate not supplied

Alternative methods of attenuating Salmonella species for potential vaccine use.

Because of their invasive properties, Salmonella species are uniquely suited for use as live oral vaccines, and mutants attenuated in general metabolic functions show promise as vaccine strains. We have identified a Salmonella choleraesuis transposon mutant which is deficient in abilities to attach to, invade or penetrate through animal cells. This mutant is avirulent in mice, yet is able to persist within the liver, spleen and gut-associated lymphoid tissue for periods which may be adequate for induction of secretory, humoral and cell-mediated immunity. This and other strains attenuated in factors specifically required for Salmonella pathogenesis may have potential as vaccine strains.

Animals↗
PubMedDate not supplied

Expression of P.69/pertactin from Bordetella pertussis in a baculovirus/insect cell expression system: protective properties of the recombinant protein.

The surface antigen P.69/pertactin of Bordetella pertussis has been expressed using the polyhedron promoter of baculovirus in cultured insect cells. Either full-length or truncated prn DNA was used to express P.69 pertactin. The full-length gene gave rise to low levels of P.93 precursor protein, some of which was processed to P.69. The shortened prn expressed P.69 pertactin directly at levels up to 3.5 mg per litre. P.69 vaccinated animals were protected against aerosol challenge with virulent B. pertussis bacteria.

Animals↗
PubMedDate not supplied

Royal Society of Tropical Medicine and Hygiene meeting at Manson House, London, 14 December 1995. Enteropathogenic Escherichia coli--mucosal infection models.

The formation of attaching and effacing (A/E) lesions is central to the pathogenesis of enteropathogenic Escherichia coli (EPEC)-mediated disease in humans and Citrobacter rodentium-mediated transmissible colonic hyperplasia in mice. Closely related outer membrane proteins, known as intimins, are required for formation of the A/E lesion by both EPEC and C. rodentium. In this study we found similar ultrastructural damage in small intestinal biopsies from an EPEC-infected child and large bowel specimens from C. rodentium-infected mice. The C. rodentium-infected large bowel biopsies revealed massive hyperplastic reactions and the infected human small intestinal biopsies showed an increase in total crypt cell number and mitotic index. EPEC-infected small intestinal organ cultures revealed bacteria adhering in a localized pattern and evidence of A/E lesions. Covaspheres coated with a biologically active cell-binding domain of intimin also adhered to cells in a localized fashion but did not induce the characteristic A/E lesions.

Adhesins, Bacterial↗
PubMedDate not supplied

Protection of chickens against experimental fowl typhoid using a nuoG mutant of Salmonella serotype Gallinarum.

A nuoG mutation in NADH dehydrogenase I was introduced into a virulent strain of Salmonella serotype Gallinarum, the causative agent of fowl typhoid, using gene replacement with a nuoG open reading frame inactivated by insertion of DNA encoding a kanamycin resistance determinant. The S. Gallinarum nuoG mutant, named SG9NGK, was highly attenuated in chickens. SG9NGK colonized the caeca of chickens less efficiently than the S. Gallinarum parental strain, was less invasive and showed no evidence of multiplication in the liver or spleen. Using a single oral immunization with live bacteria SG9NGK reduced mortality in 2-week-old chickens following challenge with virulent S. Gallinarum from 75% to less than 8%.

Animals↗