| Title |
NOTE] Bioluminescent Assay for Sphingolipid Ceramide N-Deacylase Using Vibrio harveyi Dark Mutant M-17 |
| Author |
Ki Woong Cho |
| Address |
Department of Marine Biotechnology, Anyang University, Incheon 417-833, Republic of Korea |
| Bibliography |
Journal of Microbiology, 46(5),585-589, 2008,
|
| DOI |
|
| Key Words |
bioluminescent assay, Vibrio harveyi M-17, SCDase, ceramide, myristic acid |
| Abstract |
A new bioluminescent assay method for the activity of sphingolipid ceramide N-deacylase (SCDase: EC3.5.1.69) as well as ceramidase (CDase: EC 3.5.1.23) was developed using bioluminescent marine bacteria. Enzymatically synthesized ceramide (N-myristoyl sphigosine, C14:0-18:1) and commercial SCDase were used in this demonstration, and myristic (tetradecanoic, C14:0) acid produced by the SCDase hydrolysis was quantified using Vibrio harveyi M-17, a dark mutant of V. harveyi. The in vivo light intensity of M-17 was stimulated up to thousands fold in the presence of myristic acid, was used for this assay. SCDase activity with as little as 10 μU and 5 nM of myristic acid production were detected in less than one min.
<br>The assay worked well for the determination of Km and chromatographic fraction assay. |
