RRC ID 35480
Author Simmons JM, Koslowsky DJ, Hausinger RP.
Title Characterization of a Trypanosoma brucei Alkb homolog capable of repairing alkylated DNA.
Journal Exp Parasitol
Abstract 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.
Volume 131(1)
Pages 92-100
Published 2012-5-1
DOI 10.1016/j.exppara.2012.03.011
PII S0014-4894(12)00105-1
PMID 22465611
PMC PMC3340446
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*
IF 1.719
Times Cited 4
WOS Category PARASITOLOGY
Resource
Prokaryotes E. coli JW2200(Keio collection alkB)