Construction, overexpression, and purification of Arthrobacter globiformis amine oxidase-Strep-tag II fusion protein.
The copper-containing amine oxidase from Arthrobacter globiformis has been expressed and purified as a fusion protein with a C-terminal Strep-tag II peptide. This tag facilitates the rapid purification of the enzyme on a large scale using the StrepTactin POROS medium. For example, we have demonstrated that 50 mg of protein can be obtained in 2 days from 2 L of Escherichia coli. The purified fusion protein displays turnover and spectroscopic properties that are essentially identical to those of the wild-type enzyme. Given the location of the C-terminus in four amine oxidase crystal structures, this strategy should be quite general for the rapid purification of amine oxidases from multiple sources.
Juda
G A
GA
Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT 59717, USA.
Bollinger
J A
JA
Dooley
D M
DM
D006631:Amine Oxidase (Copper-Containing) / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001173:Arthrobacter / Q000201:enzymology*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism
C099517:Ala-Trp-Arg-His-Pro-Gln-Phe-Gly-Gly; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D006631:Amine Oxidase (Copper-Containing)