A simple dot blot assay to measure hygromycin B phosphotransferase activity in whole cell extracts of Neurospora crassa

Authors

Michael Freitag, Oregon Graduate Institute of Science & Technology
Matthew S. Sachs, Oregon Graduate Institute of Science & Technology

Abstract

Hygromycin B (Hyg) is an aminocyclitol antibiotic with broad spectrum activity against prokaryotes and eukaryotes (Pettinger et al. 1953 Antibiot. Chemother. 3:1268- 1278). Hyg inhibits protein synthesis by blocking ribosomal translocation; it prevents polypeptide elongation by interfering with aminoacyl tRNA recognition and ribosomal A-site occupation (Cabanas et al. 1978 Euro. J. Biochem. 87:21-27, Hausner et al. 1988 J. Biol. Chem. 263:13103-13111). Hyg can lead to misreading during translation in vitro (Davies and Davies 1968 J. Biol. Chem. 243:3312-3316, Gonzales et al. 1978 Biochim. Biophys. Acta 521:459-469, Singh et al. 1979 Nature 277:146-148); however, this effect was not duplicated in vivo (Bakker 1992 J. Gen. Microbiol. 138:563-569).

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Recommended Citation

Freitag, M., and M.S. Sachs (1995) "A simple dot blot assay to measure hygromycin B phosphotransferase activity in whole cell extracts of Neurospora crassa," Fungal Genetics Reports: Vol. 42, Article 7. https://doi.org/10.4148/1941-4765.1337