A halophilic extracellular protease from a halophilic archaebacterium strain 172 P1

Abstract

An unidentified halophilic archaebacterium strain 172 P1 produced three extracellular proteases in media containing 15-27% salts. One component, F-II, was purified to homogeneity. It is a serine protease that can be inhibited by phenylmethylsulfonyl fluoride and chymostatin. A high concentration of NaCl was required for its stability; in the presence of 25% NaCl, only 4% of the activity was lost by incubating at 60 degrees C for 30 min, while complete inactivation occurred in the presence of 5% NaCl. F-II is a thermophilic and halophilic protease. High activity was obtained at 75-80 degrees C when F-II was assayed in the presence of 25% NaCl. The optimal concentration of NaCl required was 10-14% when assayed at 70 degrees C with azocasein as substrate, though a halophilic characteristic was not distinct at lower temperatures. Hydrolyses of the synthetic substrates succinyl-alanyl-alanyl-prolyl-phenylalanyl-4-methylcoumaryl-7-amide or succinyl-alanyl-alanyl-alanyl-p-nitroanilide at 26 degrees C were maximal at 25 and 30% NaCl, respectively. F-II was most stable at pH 6-7, and its optimal pH was 10.7. Its molecular weight was estimated as 44,000-46,000 by sodium dodecyl sulfate--polyacrylamide gel electrophoresis and by gel filtration--high-pressure liquid chromatography. The sequence of the 35 N-terminal amino acid residues was determined and compared with that of other serine proteases.