Open AccessProchymosin expression in Bacillus subtilis
Author(s) -
Parente Dino,
Ferra Francesca,
Galli Giuliano,
Grandi Guido
Publication year - 1991
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1991.tb04356.x
Subject(s) - bacillus subtilis , plasmid , complementary dna , subtilisin , microbiology and biotechnology , biology , intracellular , enzyme , cistron , gene , secretion , gene expression , protease , biochemistry , genetics , escherichia coli , bacteria
Summary Prochymosin (PC) sequence was cloned in Bacillus subtilis using two kinds of plasmid constructions. In plasmid pSM316 the cDNA was inserted to obtain the intracellular expression of the enzyme. The enzyme turned out to be expressed in an in an insoluble form which could be converted to native enzyme under proper denaturing and refolding conditions. The levels of intracellular expression of PC were further enhanced by modifying the 5′ region of the gene in a way that a two‐cistron expression system was created. For the PC secretion, the cDNA was fused to the subtilisin leader sequence and expressed under the control of the B. subtilis neutral protease promoter. A properly folded PC was secreted by the cells, although to low levels.