An electrochemiluminescence biosensor for 8-oxo-7,8-dihydro-2'-deoxyguanosine quantification and DNA repair enzyme activity analysis using a novel bifunctional probe.
A new electrochemiluminescence (ECL) sensor was developed for 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) quantification and Escherichia coli formamidopyrimidine-DNA glycosylase (FPG) activity assay. The sensor employed a novel spermine conjugated ruthenium tris-(bipyridine) derivative (spermine-Ru) which binds specifically with 8-oxodGuo through a one-step reaction and also acts as an ECL signal reporter. In the sensor, an 8-oxodGuo-containing ds-DNA film was first immobilized on a gold electrode by self-assembly. The DNA film was then incubated with spermine-Ru under oxidative condition for 8-oxodGuo labeling. The ECL intensity was found to correlate with the amount of 8-oxodGuo on the surface and the detection limit was estimated to be about 1 lesion in 500 DNA bases. Addition of FPG resulted in some loss of the signal due to the excision of 8-oxodGuo by the enzyme. An inverse relationship between ECL intensity and FPG concentration was observed in a range from 0 to 4.0U/µL, demonstrating that this sensor could be used for FPG activity assay. A number of metal ions were screened by the sensor for their inhibition effect on FPG activity. Among them, Hg(2+) and methyl Hg(II) shown very potent inhibition, with IC50 values of 4.04µM and 4.34nM respectively. The result may suggest that interference on the DNA repair system could be another mechanism for the high toxicity of MeHg.
State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China; The Education Ministry Key Lab of Resource Chemistry, Shanghai Key Laboratory of Rare Earth Functional Materials and Department of Chemistry, Shanghai Normal University, Shanghai 200234, China.
Yiping
Y
Wu
Department of Chemical Engineering, Chengde Petroleum College, Chengde, Hebei 067000, China.
Xiqiang
X
Yang
State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China.
Bintian
B
Zhang
State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China. Electronic address: LHGuo@rcees.ac.cn.
Liang-Hong
LH
Guo
D000080242:8-Hydroxy-2'-Deoxyguanosine; D015374:Biosensing Techniques / Q000295:instrumentation*; D003217:Conductometry / Q000295:instrumentation*; D045643:DNA Repair Enzymes / Q000032:analysis* / Q000737:chemistry; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000737:chemistry; D004789:Enzyme Activation; D004867:Equipment Design; D019544:Equipment Failure Analysis; D008163:Luminescent Measurements / Q000295:instrumentation*; D008628:Mercury / Q000737:chemistry; D015336:Molecular Probe Techniques / Q000295:instrumentation*
D000080242:8-Hydroxy-2'-Deoxyguanosine; D045643:DNA Repair Enzymes; D008628:Mercury; D003849:Deoxyguanosine
8-Oxo-7,8-dihydro-2′-deoxyguanosine; Electrochemiluminescence sensor; Enzyme activity assay; Formamidopyrimidine-DNA glycosylase; Mercury