DNA affinity capture and protein profiling by SELDI-TOF mass spectrometry: effect of DNA methylation.
We report here that surface enhanced laser desorption/ionization-time of flight (SELDI-TOF) mass spectrometry, as performed on a Ciphergen Biosystems ProteinChip System, can be used in conjunction with DNA affinity capture (DACA) to study specific DNA-protein binding. Using DNA molecules bound to a surface, sequence-specific interactions can be detected as demonstrated by a mutation affecting the binding profile for TBP, a transcription factor. Also, a comparison between methylated and unmethylated promoter-containing DNA fragments shows numerous binding profile differences over a mass range extending to >60 kDa. The binding of several proteins is inhibited by methylation of the DNA.
Division of Biology, Beckman Research Institute, The City of Hope National Medical Center, 1450 East Duarte Road, Duarte, CA 91010, USA.
Thomas K
TK
Bane
James F
JF
LeBlanc
Terry D
TD
Lee
Arthur D
AD
Riggs
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D002457:Cell Extracts; D018899:CpG Islands / Q000235:genetics; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated / Q000235:genetics; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D035181:TATA-Box Binding Protein; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D002457:Cell Extracts; D004268:DNA-Binding Proteins; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; C077885:XIST non-coding RNA; D004247:DNA