Molecular cloning and expression of the coagulase gene of Staphylococcus aureus 8325-4.
The gene coding for coagulase (coa) was cloned from Staphylococcus aureus 8325-4 in a lambda replacement vector in Escherichia coli. Coagulase (plasma-clotting) activity was measured in lambda coa lysates and an immunoreactive protein of 60 kDa was detected by Western immunoblotting with anti-coagulase serum. This protein comigrated with the major immunoreactive protein in supernatants of S. aureus 8325-4. The coa gene was subcloned in pUC vectors. One recombinant expressed a 60 kDa immunoreactive protein and plasma-clotting activity. A putative beta-galactosidase-coagulase fusion protein and truncated peptides were expressed by variants formed by subcloning. These results are consistent with previously published biochemical data that the prothrombin-binding domain of coagulase is located in the N-terminus of the protein. The cloned coa gene was transferred into S. aureus on a shuttle plasmid. Expression of coagulase was higher in a strain with a mutation in the agr locus, which controls the level of several exoproteins in S. aureus, suggesting that agr normally regulates coagulase expression negatively.
Phonimdaeng
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Microbiology Department, Moyne Institute, Trinity College, Dublin, Ireland.
O'Reilly
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O'Toole
P W
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Foster
T J
TJ
D002874:Chromosome Mapping; D003001:Cloning, Molecular; D003030:Coagulase / Q000235:genetics*; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D010957:Plasmids; D013211:Staphylococcus aureus / Q000201:enzymology / Q000235:genetics*
D003030:Coagulase