CLONING, PURIFICATION AND CHARACTERIZATION OF THE LIPASE FROM STAPHYLOCOCCUS-EPIDERMIDIS - COMPARISON OF THE SUBSTRATE SELECTIVITY WITH THOSE OF OTHER MICROBIAL LIPASES

On the chromosome of Staphylococcus epidermidis RP62A the lipase gene (gehSE1) is immediately flanked by the icaAA'BC operon, which is invol ved in biofilm formation. Since lipase production might play a role in staphylococcal skin colonisation as well, we studied the biochemical properties of the staphylococcal lipases more closely. The DNA sequenc e and the deduced protein sequence revealed that gehSE1 is very simila r to the lipase sequence of S. epidermidis strain 9. Like other staphy lococcal lipases, gehSE1 is organised as a preproenzyme. The part of g ehSE1 coding for the mature lipase was cloned and overexpressed as a f usion protein with an N-terminal histidine tag in Escherichia coli. Th e lipase was purified to homogeneity using a combination of precipitat ion techniques, metal-affinity chromatography and gel filtration. Bioc hemical characterisation showed that this lipase is closely related to the lipase from Staphylococcus aureus NCTC8530. Both enzymes have a p H optimum around 6, are very stable at low pH, and need calcium as a c ofactor for catalytic activity. The preferred substrates are small tri acylglycerols, with a maximum activity toward tributyrylglycerol. Comp arison of the substrate selectivity with those of other microbial lipa ses showed that phospholipids are generally poor substrates for lipase s. An exception is the lipase from Staphylococcus hyicus, which prefer s phospholipids as a substrate, distinguishing this staphylococcal lip ase from other microbial lipases. These results are discussed in view of the structure/function relationships of staphylococcal lipases, and the possible involvement of these enzymes in biological processes suc h as skin colonisation and pathogenesis.