Cloning and expression of the ponB gene, encoding penicillin-binding protein 1B of Escherichia coli, in heterologous systems.
A fragment from the ponB region of the Escherichia coli chromosome comprising the promoterless sequence encoding penicillin-binding protein 1B (PBP 1B) has been cloned in a broad-host-range expression vector under the control of the kanamycin resistance gene promoter present in the vector. The hybrid plasmid (pJP3) was used to transform appropriate strains of Salmonella typhimurium, Pseudomonas putida, and Pseudomonas aeruginosa. In all instances, the coding sequence was expressed in the heterologous hosts, yielding a product with electrophoretic mobility, protease accessibility, membrane location, and beta-lactam-binding properties identical to those of native PBP 1B in E. coli. These results indicated that PBP 1B of E. coli is compatible with the cytoplasmic membrane environment of unrelated bacterial species and support the idea that interspecific transfer of mutated alleles of genes coding for PBPs could potentially be an efficient spreading mechanism for intrinsic resistance to beta-lactams.
Plá
J
J
Instituto de Biología Molecular C.S.I.C., Centro de Biología Molecular, Universidad Autónoma, Madrid, Spain.
Rojo
F
F
de Pedro
M A
MA
Ayala
J A
JA
D000667:Ampicillin / Q000378:metabolism; D001426:Bacterial Proteins; D002352:Carrier Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005798:Genes, Bacterial; D005838:Genotype; D006602:Hexosyltransferases; D007700:Kinetics; D008854:Microscopy, Electron; D002267:Muramoylpentapeptide Carboxypeptidase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D046915:Penicillin-Binding Proteins; D010406:Penicillins / Q000378:metabolism; D010458:Peptidyl Transferases; D010957:Plasmids; D011549:Pseudomonas / Q000235:genetics; D015183:Restriction Mapping; D012486:Salmonella typhimurium / Q000235:genetics
D001426:Bacterial Proteins; D002352:Carrier Proteins; D046915:Penicillin-Binding Proteins; D010406:Penicillins; D000667:Ampicillin; D010458:Peptidyl Transferases; D006602:Hexosyltransferases; D002267:Muramoylpentapeptide Carboxypeptidase