Reference - Detail
|Author||Simmons JM, Koslowsky DJ, Hausinger RP.|
|Title||Characterization of a Trypanosoma brucei Alkb homolog capable of repairing alkylated DNA.|
Trypanosoma brucei encodes a protein (denoted TbABH) that is homologous to AlkB of Escherichia coli and AlkB homolog (ABH) proteins in other organisms, raising the possibility that trypanosomes catalyze oxidative repair of alkylation-damaged DNA. TbABH was cloned and expressed in E. coli, and the recombinant protein was purified and characterized. Incubation of anaerobic TbABH with Fe(II) and α-ketoglutarate (αKG) produces a characteristic metal-to-ligand charge-transfer chromophore, confirming its membership in the Fe(II)/αKG dioxygenase superfamily. The protein binds to DNA, with a clear preference for alkylated oligonucleotides according to results derived by electrophoretic mobility shift assays. Finally, the protozoan gene was shown to partially complement E. coli alkB cells when stressed with methylmethanesulfonate; thus confirming assignment of TbABH as a functional AlkB protein in T. brucei.
|MeSH||Alkylation Amino Acid Sequence Chromatography, Gel Cloning, Molecular DNA Repair / physiology* DNA, Protozoan / chemistry DNA, Protozoan / genetics* DNA-Binding Proteins / chemistry DNA-Binding Proteins / genetics DNA-Binding Proteins / physiology Electrophoresis, Polyacrylamide Gel Electrophoretic Mobility Shift Assay Escherichia coli Gene Expression Genetic Complementation Test Humans Molecular Sequence Data Oxidation-Reduction Protozoan Proteins / chemistry Protozoan Proteins / genetics Protozoan Proteins / physiology* Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / isolation & purification Sequence Alignment Spectrophotometry, Ultraviolet Trypanosoma brucei brucei / chemistry Trypanosoma brucei brucei / genetics*|
|Prokaryotes E. coli||JW2200(Keio collection alkB)|