TY - JOUR
T1 - Detection of choline and phosphatidic acid (PA) catalyzed by phospholipase D (PLD) using MALDI-QIT-TOF/MS with 9-aminoacridine matrix
AU - Park, Kyung Eui
AU - Kim, Jun Dal
AU - Nagashima, Yusuke
AU - Kako, Koichiro
AU - Daitoku, Hiroaki
AU - Matsui, Motoki
AU - Park, Gwi Gun
AU - Fukamizu, Akiyoshi
N1 - Publisher Copyright:
© 2014 Japan Society for Bioscience, Biotechnology, and Agrochemistry.
PY - 2014
Y1 - 2014
N2 - Phospholipase D (PLD) catalyzes the hydrolysis of phosphatidylcholine (PC), the most abundant phospholipids of plasma membrane, resulting in the production of choline and phosphatidic acid (PA). Choline is a precursor of the neurotransmitter acetylcholine, whereas PA functions as an intracellular lipid mediator of diverse biological functions. For assessing PLD activity in vitro, PLD-derived choline has been often analyzed with radioactive or nonradioactive methods. In this study, we have developed a new method for detecting choline and PA with MALDI-QIT-TOF/MS by using 9-aminoacridine as a matrix. The standard calibration curves showed that choline and PA could be detected with linearity over the range from 0.05 and 1 pmol, respectively. Importantly, this method enables the concomitant detection of choline and PA as a reaction product of PC hydrolysis by PLD2 proteins. Thus, our simple and direct method would be useful to characterize the enzymatic properties of PLD, thereby providing insight into mechanisms of PLD activation.
AB - Phospholipase D (PLD) catalyzes the hydrolysis of phosphatidylcholine (PC), the most abundant phospholipids of plasma membrane, resulting in the production of choline and phosphatidic acid (PA). Choline is a precursor of the neurotransmitter acetylcholine, whereas PA functions as an intracellular lipid mediator of diverse biological functions. For assessing PLD activity in vitro, PLD-derived choline has been often analyzed with radioactive or nonradioactive methods. In this study, we have developed a new method for detecting choline and PA with MALDI-QIT-TOF/MS by using 9-aminoacridine as a matrix. The standard calibration curves showed that choline and PA could be detected with linearity over the range from 0.05 and 1 pmol, respectively. Importantly, this method enables the concomitant detection of choline and PA as a reaction product of PC hydrolysis by PLD2 proteins. Thus, our simple and direct method would be useful to characterize the enzymatic properties of PLD, thereby providing insight into mechanisms of PLD activation.
KW - Choline
KW - MALDI-QIT-TOF/MS analysis
KW - Phosphatidic acid
KW - Phosphatidylcholine hydrolysis
KW - Phospholipase D activity
UR - http://www.scopus.com/inward/record.url?scp=84925661789&partnerID=8YFLogxK
U2 - 10.1080/09168451.2014.910102
DO - 10.1080/09168451.2014.910102
M3 - 学術論文
C2 - 25036123
AN - SCOPUS:84925661789
SN - 0916-8451
VL - 78
SP - 981
EP - 988
JO - Bioscience, biotechnology, and biochemistry
JF - Bioscience, biotechnology, and biochemistry
IS - 6
ER -