Synthetic and natural amphiphiles, octyl glucoside, Nonidet P40, sodium dodecyl sulfate (SDS), gangliosides GM1 and GD1a, interact with cholera toxin (CLT) and with its active region (promoter A). The formation of CLT-amphiphile complex leads to inhibition of ADP-ribosyltransferase activity, a chara
A radioimmune assay of ganglioside GM1 synthase using cholera toxin
โ Scribed by Koichi Honke; Naoyuki Taniguchi; Akira Makita
- Publisher
- Elsevier Science
- Year
- 1986
- Tongue
- English
- Weight
- 553 KB
- Volume
- 155
- Category
- Article
- ISSN
- 0003-2697
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โฆ Synopsis
A radioimmune assay for uridine 5'-diphosphate-galactose (UDP-Gal):GM2 galactosyltransferase, which synthesizes GM1, has been developed utilizing cholera toxin. This assay is more sensitive and simpler than previously used assays. Radioactive nucleotide substrate and GM2 were incubated with an enzyme sample, and a radiolabeled product, GM1, was reacted with cholera toxin. The GM1-cholera toxin complex was further reacted with anti-cholera toxin and Staphylococcus aureus cell suspension. The resulting complex was transferred onto a nitrocellulose membrane and quantitated by liquid scintillation counting. This assay was found to be sensitive for the detection of 100 pmol of the reaction product, GM1. With this assay method, some properties of the crude enzyme extracts from rat liver were studied. The enzyme had a pH optimum of 6.5-7.0 and required Mn2+. The Km values for UDP-Gal and GM2 were 0.12 mM and 6 microM, respectively.
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