A psychrotolerant protease-producing bacterial strain BSw20353 was isolated from the Chuckchi Sea in the Arctic,and identified as Pseudoalteromonas tetraodon is based on 16S rDNA sequencing.It could grow at temperature no higher than 38℃.The use of zymogram gels revealed the presence of three or more proteolytic bands in the culture supernatant.These bands may represent individual isozymes,different proteases,or active subunits of a larger proteolytic complex.The change of cultivation temperature affected the production of bacterial proteases,resulting in a change in protease category and quantity.One of these proteolytic bands was purified to electrophoretic homogeneity from the culture supernatant.It could be produced when bacterium growing at 15℃ and 25℃.The optimum temperature and pH for enzyme activity were 55℃ and 8,respectively.On the basis of its biochemical characteristics,the enzyme can be included in the group of metalloproteases that are inhibited by chelators including 1,10-phenanthroline,EDTA and EGTA.It was resistant to PMSF,TPCK,TLCK,and E-64,indicating that the enzyme is neither a serine protease nor cysteine protease.The protease activity was inhibited by pepstatin,an aspartic acid protease inhibitor,Hg 2+ and Sr 2+,but was stimulated by Fe 2+ and Mn 2+.SDS showed obvious inhibiting effect on the enzyme.In addition to iodoacetic acid,dithiothreitol produced a decrease of activity.However,protease activity was enhanced after treated by denaturant of urea or guanidine HCl.
