Cultivable protease-producing bacteria in the sediments of North Yellow Sea

Abstract:

[Objective] To uncover the diversity of extracellular protease-producing bacteria and to expand our knowledge on protease-producing bacteria in the sediments of North Yellow Sea, and to screen efficient protease production strains that might provide flora resources for mining marine protease-producing microorganisms.[Methods] Protease-producing bacteria were isolated by using casein gelatin plate from 5 sediment samples of the North Yellow Sea. The bacteria diversity was evaluated through phylogenetic analyses based on 16S rRNA genes. The protease diversity was evaluated through the inhibition tests for 39 strains with higher enzymatic activities by using 4 inhibitors:phenyl methyl sulfonyl fluoride (PMSF, serine protease inhibitor), 1,10-phenanthroline (O-P, metalloproteinase inhibitor), E-64 (cysteine protease inhibitor) and pepstatin A (aspartic protease inhibitor). Furthermore, degradation abilities for different protein substrates such as casein, gelatin and elastin were evaluated through observing hydrolytic zones.[Results] A total of 66 protease-producing strains were isolated from 5 sediment samples. These isolates were classified into 7 genera of 4 phyla including Bacteroidetes, Proteobacteria, Actinobacteria and Firmicutes, with Pseudoalteromonas (69.9%), Sulfitobacter (12.1%) and Salegentibacter (10.6%) as the dominant. The richness of the cultivable protease-producing bacteria reached 10⁴ cells/g in all sediment samples. The inhibition tests indicated that all the tested strains produced serine protease and/or metal protease, only a few strains produced cysteine protease or aspartic protease.[Conclusion] The cultivable protease-producing bacteria in the North Yellow Sea are diverse with Pseudoalteromonas, Sulfitobacter and Salegentibacterbacteria as the dominant groups and the extracellular proteases belong to serine proteases and/or metalloproteinases.