Cloning and expression of the gene for a fibronectin-binding protein from Staphylococcus aureus.
The gene encoding the fibronectin-binding protein (FNBP) from Staphylococcus aureus strain 8325-4 was isolated from a gene bank in pBR322. The original clone, containing a 6.5-kb insert, gave a functional product present in the periplasm of Escherichia coli. Analysis of polypeptides isolated after affinity chromatography on fibronectin-Sepharose followed by ion-exchange chromatography revealed two gene products, 87 and 165 kd in mol. wt. The amino acid compositions of these two polypeptides and a native FNBP from S. aureus strain Newman were very similar. Antibodies raised against the native FNBP from strain Newman precipitated the 125I-labelled 165-kd polypeptide, and unlabeled 165- and 87-kd polypeptides as well as native FNBP inhibited the immunoprecipitation reactions. The region of the fnbp-gene encoding the fibronectin-binding activity has been identified and subcloned in an expression vector based on the staphylococcal protein A gene. The resulting product in E. coli is an extracellular fusion protein consisting of two IgG-binding domains of protein A followed by a fibronectin-binding region. The fusion protein binds to fibronectin and completely inhibits the binding of fibronectin to intact cells of S. aureus.
Flock
J I
JI
Department of Microbiology, University of Uppsala, Sweden.
Fröman
G
G
Jönsson
K
K
Guss
B
B
Signäs
C
C
Nilsson
B
B
Raucci
G
G
Höök
M
M
Wadström
T
T
Lindberg
M
M
D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics; D005353:Fibronectins / Q000378:metabolism; D005796:Genes; D005798:Genes, Bacterial; D008970:Molecular Weight; D010957:Plasmids; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic / Q000235:genetics* / Q000378:metabolism; D013211:Staphylococcus aureus / Q000235:genetics*; D014158:Transcription, Genetic
D005353:Fibronectins; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic; D004262:DNA Restriction Enzymes