A chromogenic substrate for solid-phase detection of phospholipase A(2)

作者：Eba Chisato; Okano Aoi; Nakano Hideo; Iwasaki Yugo^*

来源：Analytical Biochemistry, 2014, 447: 43-45.

DOI：10.1016/j.ab.2013.11.007

• Phospholipase A(2)
• Solid-phase detection
• Diazo coupling
• Artificial substrate
• High-throughput screening

A method for solid-phase detection of phospholipase A(2) (PLA(2)) was developed. The method uses 1-octanoyloxynaphthalene-3-sulfonic acid, which was found to be a good substrate of PLA(2). The substrate is hydrolyzed by PLA(2) into 1-naphthol-3-sulfonic acid, which is spontaneously coupled with coexisting diazonium salt to form a red-purple azo dye. Streptomyces and bovine pancreatic PLA(2) spotted on a nitrocellulose membrane could be detected by this method with considerable sensitivity. In addition, colonies of recombinant Escherichia coli producing bacterial PLA(2) were distinguishable from those producing an inactive mutant PLA(2), facilitating high-throughput screening in directed evolution of the enzyme.

• 出版日期2014-2-15