Magnetic DNA affinity purification of yeast transcription factor tau--a new purification principle for the ultrarapid isolation of near homogeneous factor.
We present a new method for rapid purification to near homogeneity of sequence specific DNA binding proteins based on magnetic separation. The method is described for the purification of the yeast transcription factor tau. DNA affinity Dynabeads (monodisperse superparamagnetic particles) specifically bind the protein in the presence of competitor DNA. By magnetic separation, wash and elution, highly enriched transcription factor preparations are obtained within minutes. In less than an hour with three cycles of adsorption, nearly homogeneous factor tau was obtained. The factor preparation contained mainly two polypeptides of 100 and 140 kDa and was fully active in transcription and DNA binding assays. This procedure should work for any high-affinity sequence-specific DNA binding protein with only minor modifications.
Gabrielsen
O S
OS
Department of Biochemistry, University of Oslo, Norway.
Hornes
E
E
Korsnes
L
L
Ruet
A
A
Oyen
T B
TB
D000327:Adsorption; D001426:Bacterial Proteins; D001710:Biotin; D004271:DNA, Fungal; D003850:Deoxyribonuclease I; D005800:Genes, Fungal; D007700:Kinetics; D008280:Magnetics; D012352:RNA, Transfer, Glu / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000032:analysis*; D019809:Streptavidin; D014157:Transcription Factors / Q000302:isolation & purification* / Q000378:metabolism; D019474:Transcription Factors, TFIII; D014158:Transcription, Genetic; D000098832:Transcription Factor TFIIIC
D001426:Bacterial Proteins; D004271:DNA, Fungal; D012352:RNA, Transfer, Glu; D014157:Transcription Factors; D019474:Transcription Factors, TFIII; D000098832:Transcription Factor TFIIIC; D001710:Biotin; D019809:Streptavidin; D003850:Deoxyribonuclease I