PCR-cloning of the pac gene from E-coli ATCC11105 - Observations on the expression of a recombinant periplasmic protein

The penicillin acylase gene (pac) amplified by polymerase chain reaction (PCR) from Escherichia coli (E. coli) ATCC11105 genomic DNA was cloned into pUC9, pEMBL9(+) and pCP40 vectors. A penicillin acylase precursor was overexpressed at 26 degrees C from pac-pEMBL9(+) and pac-pCP40 constructs transformed into E.coli strains JM109 and pCI857 containing HB101, respectively. With the thermo-inducible pac-pCP40 construct some level of mature alpha and beta subunits and varying degrees of enzyme activity were also detected.