Method for determining substrate specificity of protease

摘要

The present invention relates to a method for determining substrate specificity of a protease by employing a microorganism cotransformed with a vector expressing the protease and a vector expressing the substrate. The method comprises constructing an expression vector containing a gene for protease; constructing an expression vector containing genes for Golgi recognition signal and transmembrane domain of membrane protein locating at yeast Golgi complex, substrate domain with specific amino acid sequence, and yeast invertase; preparing a transformant by cotransforming suc2 mutant yeast with the two expression vectors; and, determining viability of the transformant in a medium containing a sole carbon source of sucrose. The invented method requires neither isolation/purification of protease nor synthesis of expensive substrate peptide. The simple and cost-effective determination of substrate specificity of proteases can be realized to allow its wide application in the identification of genes for proteases or genes for proteins cleaved with the proteases.