PRIMARY STRUCTURE AND BIOLOGICAL FEATURES OF A THERMOSTABLE NUCLEASE ISOLATED FROM STAPHYLOCOCCUS-HYICUS

The nucH gene, encoding a thermostable nuclease (TNase), was isolated from the cellular DNA of Staphylococcus hyicus strain E80 and sequence d. NucH, the 169-amino-acid (aa) protein encoded by this gene, contain s, at its N-terminus, a signal peptide which appears to be cleaved at the same site in S. hyicus and Escherichia coli, yielding a mature pro tein which is exported extracellularly from S. hyicus, but not from E. coli. The aa sequence of NucH is highly homologous with that of the T Nase from S. intermedius strain LRA076, whereas significant similariti es are observed with the TNase from S. aureus, as well as with three o ther bacterial proteins of which only one has been shown to exhibit DN ase activity. As seen in a multiple sequence alignment, the invariant residues are mostly located in the regions involved in the biological activity of the S. aureus TNase. The ability of crude cell extracts of E. coli strains carrying nucH to degrade various forms of nucleic aci ds with or without Ca2+ supplementation was studied. Under our experim ental conditions, the enzyme encoded by nucH was active at 37 degrees C on both DNA and RNA, had the potential to act as an endonuclease, an d functioned in the presence of Ca2+. Moreover, activity was retained after heating at 100 degrees C, suggesting that the enzyme could under go reversible unfolding.