One-step magnetic purification of recombinant DNA-binding proteins using magnetizable phosphocellulose.
Magnetizable solid phase technology was used to develop a method for the rapid purification of recombinant proteins expressed in Escherichia coli. We describe the purification of two recombinant DNA-binding proteins: the minimal DNA-binding domain of the oncoprotein Myb and full-length yeast TFIIIA. Both were purified in one step directly from an E. coli lysate by means of magnetizable phosphocellulose particles (PhosphoMagnaCel). All operations were performed in microcentrifuge tubes and could be completed within 15 min. High purity and excellent recovery of proteins active in sequence specific DNA-binding were obtained. The procedure allowed the simultaneous purification of eight mutant Myb-proteins within 30 min.
Risøen
P A
PA
Department of Biochemistry, University of Oslo, Norway.
Struksnes
K
K
Myrset
A H
AH
Gabrielsen
O S
OS
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002482:Cellulose / Q000031:analogs & derivatives; D002645:Chickens; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics; D008280:Magnetics; D008969:Molecular Sequence Data; D009154:Mutation; D020626:Oncogene Proteins v-myb; D011994:Recombinant Proteins / Q000302:isolation & purification; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics / Q000302:isolation & purification*; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification*; D015003:Yeasts
D004268:DNA-Binding Proteins; D020626:Oncogene Proteins v-myb; D011994:Recombinant Proteins; D015682:Retroviridae Proteins, Oncogenic; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors; D002482:Cellulose; C008881:phosphocellulose