Isolation, Screening, Partial Purification and Characterization of Protease From Halophilic Bacteria Isolated From Indonesian Fermented Food

Abstract

The protease producing bacteria were screened from Indonesian traditional fermented food, tauco and terasi, and 4 halophilic protease producers were isolated. Among these isolates, halophilic bacterial isolate TANN 4 was recorded as the best protease producer. Extracellular protease from isolate TANN 4 was partially purified using ammonium sulfate precipitation. The protease was partially purified with final yield of 72.87 % and 25.41 fold purity. This moderate thermoactive and alkaliphilic protease showed a pH optimum of 8.0 and temperature optimum was 50 °C. The enzyme was also active at salt concentrations ranging from 1 to 15 % (w/v), with optimum activity at 1 % NaCl (w/v). Ethylenediaminetetraacetic acid (EDTA) completely inhibited the enzyme activity suggesting that it was a metalloprotease. Among metal ions, the Ca 2+ , K + and Mg 2+ ions enhanced the activity of enzyme. The KM and Vmax values exhibited by partially purified protease were 0.0649 mM and 216.45 U mg −1 using casein as substrate. The molecular weight was estimated to be 19.8 kDa on SDS PAGE. The enzyme was also fairly stable in Triton X-100, SDS, 1 % commercial detergents (OMO and Ariel) and 25 % methanol and it was capable of hydrolyzing casein, hemoglobin and bovine serum albumin (BSA). These characteristics make this halophilic bacterial extracellular metalloprotease seem to be potentially useful for biotechnological and industrial applications. Automated ribotyping analysis revealed that 3 isolates (TANN 4, TR 2 and TR 4) resembled Halobacillus trueperi that exhibited 71, 68 and 69 % similarity respectively, and isolate (TR 1) resembled Virgibacillus pantothenticus with 64 % similarity.

Low molecular weight peptides in tauco, a fermented soy product, associated to umami taste through peptidomics‐sensomics approach

“…Tauco is produced by koji fermentation with mould starters, followed by brine fermentation at relatively high concentrations (Nandiyanto et al, 2018). During tauco fermentation, the soybean proteins are digested into amino acids and short-chain peptides because of the production of proteases during the growth of moulds and halophilic bacteria, as well as yeasts (Fitriani & G€ uven, 2018). Most of the salt fermentation of food protein sources results in an umami taste, which is found in tauco.…”