Genetic system for analyzing Escherichia coli thymidylate synthase
نویسندگان
چکیده
منابع مشابه
Title Efficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
l-Red system-based recombinogenic engineering is a powerful new method to engineer DNA without the need for restriction enzymes or ligases. Here, we report the use of a single selectable marker to enhance theusefulness of thisapproach.Thestrategy is to utilize the thymidylate synthase A (thyA) gene, which encodes an enzyme involved in the synthesis of thymidine 50-triphosphate, for both positiv...
متن کاملEfficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
Lambda-red system-based recombinogenic engineering is a powerful new method to engineer DNA without the need for restriction enzymes or ligases. Here, we report the use of a single selectable marker to enhance the usefulness of this approach. The strategy is to utilize the thymidylate synthase A (thyA) gene, which encodes an enzyme involved in the synthesis of thymidine 5'-triphosphate, for bot...
متن کاملModeling functional changes to Escherichia coli thymidylate synthase upon single residue replacements: a structure-based approach
Escherichia coli thymidylate synthase (TS) is an enzyme that is indispensable to DNA synthesis and cell division, as it provides the only de novo source of dTMP by catalyzing the reductive methylation of dUMP, thus making it a key target for chemotherapeutic agents. High resolution X-ray crystallographic structures are available for TS and, owing to its relatively small size, successful experim...
متن کاملEscherichia coli thymidylate synthase: amino acid substitutions by suppression of amber nonsense mutations.
By using site-directed oligonucleotide mutagenesis, amber nonsense stop codons (5'-TAG-3') have been introduced at 20 sites in the Escherichia coli thymidylate synthase gene. By transforming the thyA mutant plasmids into 13 strains, each of which harbor different amber suppressor tRNAs, we were able to generate over 245 amino acid substitutions in E. coli thymidylate synthase (EC 2.1.1.45). Gro...
متن کاملDrug-resistant variants of Escherichia coli thymidylate synthase: effects of substitutions at Pro-254.
Drug-resistant variants of thymidylate synthase (TS) can potentially be used in gene therapy applications to decrease the myelosuppressive side effects of TS-directed anticancer agents or to select genetically modified cells in vivo. Mutations of proline 303 of human TS confer resistance to TS-directed fluoropyrimidines and antifolates (). We generated the corresponding variants in Escherichia ...
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ژورنال
عنوان ژورنال: Journal of Bacteriology
سال: 1984
ISSN: 0021-9193,1098-5530
DOI: 10.1128/jb.160.1.371-378.1984