Inhibition of yeast inositol phosphorylceramide synthase by aureobasidin A measured by a fluorometric assay

FEBS Letters

Zhong, Wenyan¹  Murphy, Dennis J¹  Georgopapadakou, Nafsika H¹ 

[1] Department of Antimicrobial Research, DuPont Pharmaceuticals Company, Experimental Station, Bldg. E400, P.O. Box 80400, Wilmington, DE 19880-0400, USA

关键词: Fungal sphingolipid;    Fluorometric assay;    Inositol phosphorylceramide synthase;    Aureobasidin A;    6-[N-(7-Nitro-2;    1;    3-benzoxadiazol-4-yl)amino]-hexanoyl ceramide;    IPC;    inositol phosphorylceramide;    PI;    phosphatidylinositol;    C6-NBD-cer;    6-[N-(7-nitro-2;    1;    3-benzoxadiazol-4-yl)amino]-hexanoyl ceramide;    DMSO;    dimethylsulfoxide;   

DOI  :  10.1016/S0014-5793(99)01633-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Inositol phosphorylceramide synthase (IPC synthase) is an essential and unique enzyme in fungal sphingolipid biosynthesis and is the target of the cyclic nonadepsipeptide antibiotic aureobasidin A. As a first step towards understanding the mechanism of aureobasidin A inhibition, we developed a fluorometric HPLC assay for IPC synthase using the Saccharomyces cerevisiae enzyme and the fluorescent substrate analog 6-[N-(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]-hexanoyl ceramide (C₆-NBD-cer). The kinetic parameters for C₆-NBD-cer were comparable to those for the synthetic substrate N-acetylsphinganine used previously. Aureobasidin A acted as a tight-binding, non-competitive inhibitor with respect to C₆-NBD-cer and had a K _i of 0.55 nM.

【 授权许可】

Unknown