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.
Gümüşel
F
F
Department for Protein Structure and Function Relationships, TUBITAK Marmara Research Center, Gebze-Kocaeli, Turkey.
Kiremit
N
N
Işsever
S
S
Oğras
T T
TT
Bermek
E
E
D001483:Base Sequence; D003001:Cloning, Molecular; D004790:Enzyme Induction; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010399:Penicillin Amidase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry; D013696:Temperature
D011993:Recombinant Fusion Proteins; D010399:Penicillin Amidase