One-step purification of phosphinothricin acetyltransferase using reactive dye-affinity chromatography.
Reactive dye purification is an affinity purification technique offering unique selectivity and high purification potential. Historically, purification of phosphinothricin acetyltransferase (PAT) has involved several steps of precipitation and column chromatography. Here, we describe a novel purification method that is simple, time-saving, inexpensive, and reproducible. The novel method employs a single chromatography step using a reactive dye resin, Reactive brown 10-agarose. Reactive brown 10 preferentially binds the PAT protein, which can then be specifically released by one of its substrates, acetyl-CoA. Using Reactive brown 10-agarose, PAT protein can be purified to homogeneity from E. coli or plant tissue with high recovery efficiency.
Monsanto Company, 800 North Lindbergh Blvd, St. Louis, MO, 63167, USA, cunxi.wang@monsanto.com.
Cunxi
C
Wang
Thomas C
TC
Lee
Kathleen S
KS
Crowley
Erin
E
Bell
D000123:Acetyltransferases / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000191:economics / Q000379:methods*; D004396:Coloring Agents / Q000737:chemistry*; D004926:Escherichia coli / Q000201:enzymology; D030821:Plants, Genetically Modified; D012685:Sepharose / Q000737:chemistry; D013997:Time Factors; D014227:Triazines / Q000737:chemistry*
D004396:Coloring Agents; D014227:Triazines; C053802:Procion Brown MX 5BR; D012685:Sepharose; D000123:Acetyltransferases; C056243:phosphinothricin N-acetyltransferase