Exploration and characterization of proteases produced by halophilic microbes isolated from salt-concentrating pond at Pasuruan, East Java

Protease is one of the industrial enzymes which is capable of hydrolyzing peptide bonds in proteins. Many industries often require protease that is tolerant at high salt concentration. Indonesia is a country that has high microbial biodiversity, including halophiles, which naturally live in high salt environments such as salt concentrating ponds. We investigated proteases produced by halophilic microbes isolated from salt concentrating pond at Pasuruan, East Java. This research was conducted in several stages, namely isolation and selection of bacteria, gram staining and homogeneity tests, the stability of isolated bacteria at various NaCl concentrations, determination of optimal protease production time, characterization of proteases by pH, temperature, and the effect of salt concentration. The result shows that six isolates were obtained from salt concentrating pond at the highest concentration of 8.4%. No colonies grew from saltwater at higher salt concentrations. All colonies were indicated as rod-shaped gram-negative bacteria. Two of the isolates, namely TG-6 and TG-3, were selected for further investigation due to their high proteolytic index. The selected isolates show the highest production time of 3 days at 43 °C, optimum pH of 8 and 9, and crude extract activity of 2.475 and 4.809 U/mL, respectively. They grow at a moderate salt concentration (up to 10% of NaCl), and the enzyme has the potential for industrial application.