Involvement of NH₂-Terminal Pro-sequence in the Production of Active Aqualysin I (a Thermophilic Serine Protease) in Escherichia coli

抄録

Aqualysin I is a heat-stable subtilisin-type protease produced by Thermus aquaticus YT-1. The precursor of aqualysin I consists of four domains: an NH₂-terminal signal peptide, an NH₂-terminal pro-sequence, a protease domain, and a COOH-terminal pro-sequence. In Escherichia coli cells harboring recombinant plasmid carrying the aqualysin I gene, proteolytic activity is obtained on treatment at 65°C and mature enzyme is detected. In the case of mutant genes containing partial deletions in the NH₂-terminal pro-sequence, no proteolytic activity was detected and the precursor protein was found to be unstable in E. coli. These results indicate that the NH₂-terminal pro-sequence is required to produce the active enzyme by stabilizing the precursor structure. Amino acid substitutions in the conserved sequence of the NH₂-terminal pro-sequence found among subtilisin-type proteases made the processing faster compared with the wild type.