Detection of choline and phosphatidic acid (PA) catalyzed by phospholipase D (PLD) using MALDI-QIT-TOF/MS with 9-aminoacridine matrix

Kyung Eui Park, Jun Dal Kim, Yusuke Nagashima, Koichiro Kako, Hiroaki Daitoku, Motoki Matsui, Gwi Gun Park, Akiyoshi Fukamizu*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

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.

Original languageEnglish
Pages (from-to)981-988
Number of pages8
JournalBioscience, biotechnology, and biochemistry
Volume78
Issue number6
DOIs
StatePublished - 2014

Keywords

  • Choline
  • MALDI-QIT-TOF/MS analysis
  • Phosphatidic acid
  • Phosphatidylcholine hydrolysis
  • Phospholipase D activity

ASJC Scopus subject areas

  • General Medicine

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