Journal of Microbiology

Journal Article

NOTE] Bioluminescent Assay for Sphingolipid Ceramide N-Deacylase Using Vibrio harveyi Dark Mutant M-17: Ki Woong Cho; J. Microbiol. 2008;46(5):585-589. Published online October 31, 2008; DOI: https://doi.org/10.1007/s12275-008-0114-2

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.

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