Fj. Enguita et al., AN INDUCIBLE EXPRESSION SYSTEM OF HISTIDINE-TAGGED PROTEINS IN STREPTOMYCES-LIVIDANS FOR ONE-STEP PURIFICATION BY NI2-CHROMATOGRAPHY( AFFINITY), FEMS microbiology letters, 137(2-3), 1996, pp. 135-140
An expression and purification cassette containing the aminoglycoside
phosphotransferase gene (aph) as selective marker has been constructed
in the Escherichia coli vector pULHis2. DNA fragments inserted in the
cassette can be easily subcloned in pIJ699 to give vectors for overex
pression of genes in Streptomyces and purification of proteins by a on
e-step procedure. The expression system uses the thiostrepton-inducibl
e promoter tipA for expression and a six histidine coding nucleotide s
equence that is fused in frame to the foreign gene inserted in the pol
ylinker. The pULHis2-derived expression vector has been used satisfact
orily to express and to purify the P-7 and P-8 proteins of Nocardia la
ctamdurans which carry our the methoxylation of cephalosporin C to 7-m
ethoxycephalosporin C.