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Biomedical subjects

I D Hickson

Publications and source records attributed to I D Hickson.

135 records · Page 8Linked to original sources
PubMed1985

Isolation and characterization of Chinese hamster ovary cell lines sensitive to mitomycin C and bleomycin.

Seven Chinese hamster ovary K1 cell lines exhibiting sensitivity to anticancer drugs have been isolated by a replica-plating technique. Five of the mutants are hypersensitive (approximately 7-fold, as judged by D37 values) to the DNA cross-linking agent mitomycin C. Of these, one is also appreciably sensitive to UV light. Significant variations in their cross-sensitivity to cis-platinum(II) diammine dichloride, chlorambucil, and Adriamycin have also been observed. Two additional mutants have been isolated on the basis of sensitivity to the radiomimetic agent bleomycin. One of these shows greater than 6-fold sensitivity to bleomycin, while the other is approximately 14 times more sensitive than the parental strain to bleomycin and is also hypersensitive to a number of other DNA-damaging agents, including cis-platinum(II) diammine dichloride, chlorambucil, X-rays, and UV light. Both bleomycin-sensitive mutants also exhibit some degree of sensitivity to Adriamycin. In all cases, the cell lines have been grown in continuous culture for 3 months without evidence of reversion and should act as suitable recipients in DNA transfection experiments aimed at identifying human DNA repair genes.

Animals↗
PubMed1984

Molecular amplification and purification of the E. coli recC gene product.

The level of recC gene expression has been analysed using Mud(bla lac) fusions to the recC promoter. The constitutive level of expression is very low and remains so even under SOS inducing conditions. The recC gene product has been amplified by harnessing the gene to the phage lambda leftward promoter in a plasmid. From cells harbouring this plasmid, RecC protein, which represented approximately 6% of the total cellular protein, was purified to electrophoretic homogeneity.

Bacteriophage lambda↗
PubMed1983

The E. coli uvrD gene product is DNA helicase II.

We have shown that the uvrD gene product, previously identified in maxicell extracts as a 73 kilodalton protein, copurifies with single stranded DNA-dependent ATPase and ATP-dependent DNA helicase activities. This protein is specifically precipitated from maxicell extracts by antibodies raised against DNA helicase II. In order to facilitate purification of the UvrD protein we have subcloned the uvrD gene into a plasmid vector in which its transcription is under the control of the phage lambda leftward promoter. Using cells harbouring this recombinant plasmid as a source of elevated levels of the UvrD protein we have purified this protein to homogeneity by a simple, rapid procedure. The purified protein has single stranded DNA-dependent ATPase activity and ATP-dependent DNA helicase activity, and both activities are specifically inactivated by antibodies raised against DNA helicase II. We conclude that DNA helicase II is the uvrD gene product.

Adenosine Triphosphatases↗
PubMed1982

Construction of recombinant lambda phages that carry the E. coli recB and recC genes.

A fragment of the E. coli chromosome including the recC gene has been cloned by in vitro recombinant DNA techniques into a phage lambda vector to give the recombinant phage lambda drecC. This was used to derive the phage lambda drecBC by in vivo recombination. On lysogenisation of recB and recC mutants with lambda drecBC wild levels of UV-resistance and RecBC DNase activity were restored. Infection of E coli with lambda drecBC led to the synthesis of phage-coded proteins of 125 kilodaltons (kd) and 135 kd that were not synthesised on infection with the original lambda vector, whereas a 125 kd protein but not a 135 kd protein was synthesised in similar experiments with lambda drecC. The recombinant phages, which are unable to form plaques, presumably due to the deletion of essential phage genes during their construction, provided useful starting points from which to subclone the recB, recC, and the neighbouring thyA and argA genes individually into multiple copy plasmid vectors.

Bacterial Proteins↗
PubMed1982

Molecular cloning and amplification of the gene for thymidylate synthetase of E. coli.

The thyA gene of Escherichia coli, which directs the synthesis of the enzyme thymidylate synthetase, has been subcloned from a recombinant lambda phage (Hickson et al., 1982) into the multicopy plasmid pBR325 to give the plasmid pPE245. To identify the thyA gene product, the transposon Tn1000 was inserted into pPE245 and derivative plasmids isolated that were no longer able to complement thyA mutations. When proteins synthesised by these plasmids and by pPE245 were labelled and analysed on SDS-polyacrylamide gels a protein of 33000 Mr, presumably the thyA+ gene product was absent whenever the thyA gene was inactivated. On assaying cell extracts prepared from cells harbouring pPE245 for thymidylate synthetase, the level of this enzyme was found to be elevated by a factor of at least 25.

Bacteriophage lambda↗
PubMed1981

A temperature sensitive Reca protein of Escherichia coli.

The temperature sensitive allele recA200 has been cloned into the multiple copy number plasmid pBR322 and the gene product isolated. The purified RecA200 protein is temperature sensitive in ability to cleave the phage lambda and LexA repressors in vitro and also in ability to promote a successful search for homology between single stranded DNA and a homologous duplex leading to D-loop formation. However, at the non-permissive temperature the RecA200 protein has approximately wild type single stranded DNA dependent ATPase activity and ability to promote pairing between homologous single DNA strands. The demonstration that the temperature sensitivity in vivo can be correlated with the temperature sensitive cleavage of the lambda and LexA repressors in vitro and also with D-loop formation shows that these in vitro reactions, which require large amounts of RecA protein, are not carried out by trace amounts of contaminating proteins.

Bacterial Proteins↗