Isolation of a chymotrypsinogen B-like enzyme from the archaeon Natronomonas pharaonis and other halobacteria

Abstract

A protease of a molecular mass of approximately 30kDa was isolated and purified from the haloalkaliphilic archaeon Natronomonas (formerly Natronobacterium) pharaonis. The enzyme hydrolyzed synthetic peptides, preferentially at the carboxyl terminus of phenylalanine or leucine, as well as large proteins. Hydrolysis occurred over the range of pH from 6 to 12, with an optimum at pH 10. The temperature optimum was 61 degrees C. The enzyme was nearly equally active over the range of salt concentration from 0.5 to 4M (NaCl or KCl). A strong cross-reaction with a polyclonal antiserum against human chymotrypsin was observed. Enzymatic activity was inhibited by typical serine protease inhibitors. There was significant homology between N-terminal and internal sequences from autolytic fragments and the sequence of bovine chymotrypsinogen B; the overall amino acid composition was similar to that of vertebrate chymotrypsinogens. Evidence for a zymogen-like processing of the protease was obtained. Cell extracts from other halobacteria exhibited similar proteolytic activity and immunoreactivity. The data suggested a widespread distribution of a chymotrypsinogen B-like protease among halo- and haloalkaliphilic Archaea.

Living with two extremes: Conclusions from the genome sequence of Natronomonas pharaonis

“…All 15 previously published N. pharaonis genes could be identified within the genome, although minor sequence variations were detected usually owing to strain differences. However, a partially sequenced protease from N. pharaonis, which is extremely similar to vertebrate chymotrypsin (Stan-Lotter et al 1999), could not be found.…”

Production of Extracellular Halo-alkaline Protease from a Newly Isolated Haloalkaliphilic Bacillus sp. Isolated from Seawater in Western India

“…In comparison, haloalkaliphilic bacteria have been relatively less attended, as only few alkaline proteases are reported from these organisms (Studdert et al 1997;Stan-Lotter et al 1999;Gimenez et al 2000;Studdert et al 2001;Polosina et al 2002). Besides the novel catalytic applications, the wide occurrence of many of such enzymes among the haloalkaliphilic bacteria holds ecological significance.…”

Purification and biological characterization of a halophilic thermostable protease from Haloferax lucentensis VKMM 007

“…Protease production has been shown in several species of halophilic archaea including Halobacterium salinarum (Izotova et al 1983;Ryu et al 1994;Kim and Dordick 1997); Natrialba asiatica (Kamekura and Seno 1990;Kamekura et al 1992), Haloferax mediterranei (Stepanov et al 1992;Kamekura et al 1996;Nolasco et al 2002); Natronomonas pharaonis (Stan-Lotter et al 1999); Natronococcus occultus (Studdert et al 1997(Studdert et al , 2001; Natrialba magadii (Giménez et al 2000), Halogeometricum borinquense (Vidyasagar et al 2006) as well as in Halobacterium sp. SP1 (1) (Akolkar et al 2008) and Natrinema sp.…”