TY - JOUR
T1 - Enzymatic characterization of an amine oxidase from arthrobacter sp. used to measure phosphatidylethanolamine
AU - Ota, Hiroko
AU - Tamezane, Hideto
AU - Sasano, Yoshie
AU - Hokazono, Eisaku
AU - Yasuda, Yuko
AU - Sakasegawa, Shin Ichi
AU - Imamura, Shigeyuki
AU - Tamura, Tomohiro
AU - Osawa, Susumu
PY - 2008
Y1 - 2008
N2 - Ethanolamine oxidase was screened with the aim of using it to establish a novel enzymatic phosphatidylethanolamine assay. Ethanolamine oxidase activity was detected in the crude extract of Arthrobacter sp., and the enzyme was purified more than 15-fold in three steps with a 54% yield. SDS-PAGE revealed the presence of only one band, which migrated, with an apparent molecular mass of 70 kDa. Biochemical characterization of the enzyme showed phenylethylamine to be the preferred substrate, with the highest kcat/Km value. The primary structure, determined by sequencing the cloned gene, showed a high degree of identity to Cu-containing phenylethylamine oxidase (64%). When heterologously overexpressed in Escherichia coli, the enzyme exhibited only a trace of amine oxidase activity, but high levels of activity emerged after exposure to Cu 2+, as is typical of recombinant copper amine oxidases. Preliminary application of this enzyme coupled with phospholipase D for determination of phosphatidylethanolamine is also described. This is the first enzymatic method for the measurement of phosphatidylethanolamine.
AB - Ethanolamine oxidase was screened with the aim of using it to establish a novel enzymatic phosphatidylethanolamine assay. Ethanolamine oxidase activity was detected in the crude extract of Arthrobacter sp., and the enzyme was purified more than 15-fold in three steps with a 54% yield. SDS-PAGE revealed the presence of only one band, which migrated, with an apparent molecular mass of 70 kDa. Biochemical characterization of the enzyme showed phenylethylamine to be the preferred substrate, with the highest kcat/Km value. The primary structure, determined by sequencing the cloned gene, showed a high degree of identity to Cu-containing phenylethylamine oxidase (64%). When heterologously overexpressed in Escherichia coli, the enzyme exhibited only a trace of amine oxidase activity, but high levels of activity emerged after exposure to Cu 2+, as is typical of recombinant copper amine oxidases. Preliminary application of this enzyme coupled with phospholipase D for determination of phosphatidylethanolamine is also described. This is the first enzymatic method for the measurement of phosphatidylethanolamine.
UR - http://www.scopus.com/inward/record.url?scp=55049086888&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=55049086888&partnerID=8YFLogxK
U2 - 10.1271/bbb.80365
DO - 10.1271/bbb.80365
M3 - Article
C2 - 18838796
AN - SCOPUS:55049086888
SN - 0916-8451
VL - 72
SP - 2732
EP - 2738
JO - Bioscience, Biotechnology and Biochemistry
JF - Bioscience, Biotechnology and Biochemistry
IS - 10
ER -