A Spectrophotometric Assay for the Transphosphatidylation Activity of Phospholipase D Enzyme

✍ Scribed by Tairo Hagishita; Masanobu Nishikawa; Tadashi Hatanaka

• Publisher: Elsevier Science
• Year: 1999
• Tongue: English
• Weight: 77 KB
• Volume: 276
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1999.4353

✦ Synopsis

We developed a specific spectrophotometric assay for the quantitative determination of phospholipase D-catalyzed transphosphatidylation activity. The assay measures p-nitrophenol liberated by phospholipase D-catalyzed reaction of phosphatidyl-p-nitrophenol and ethanol in an aqueous-organic emulsion system. The release of p-nitrophenol was linear to reaction time at an early stage of the reaction with phospholipase D from Streptomyces sp. In the spectrophotometric assay for the reaction with phospholipase D from Streptomyces chromofuscus, which has higher hydrolytic activity than transphosphatidylation activity, p-nitrophenol was not found. The advantages of this novel method for measuring the transphosphatidylation activity of phospholipase D are that (i) it does not use radioactive compounds, (ii) it can measure the initial velocity of the reaction, and (iii) it is rapid, easy, and accurate to perform.