AN INDUCIBLE EXPRESSION SYSTEM OF HISTIDINE-TAGGED PROTEINS IN STREPTOMYCES-LIVIDANS FOR ONE-STEP PURIFICATION BY NI2-CHROMATOGRAPHY( AFFINITY)

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.