Isolation and characterization of a novel oxidant‐ and surfactant‐stable extracellular alkaline protease from Exiguobacterium profundumBK‐P23

Abstract

The novel protease from Exiguobacterium profundum BK-P23 was partially purified by ammonium sulfate precipitation and further purified Mono Q 5/50 and Superdex 200 10/300 column chromatography. The enzyme was purified 10.23-fold with a yield of 14%. The molecular weight was estimated to be 52 kDa by SDS-PAGE. The enzyme was most active at a pH of 8.0 and temperature of 40°C and the enzyme was stable between a pH of 7 and 10 and up to a temperature of 50°C. The enzyme activity was enhanced by CaCl2 but was slightly inhibited by CoCl2 , MgSO4 , and AgNO3 . In addition, this enzyme was completely inhibited by phenylmethylsulfonyl fluoride, indicating that this enzyme was a serine protease. Furthermore, the alkaline protease was more stable in the presence of surfactants such as Triton X-100, which was followed by Tween 80 and SDS. Moreover, the enzyme was highly stable in the presence of 1% oxidizing agent (H2 O2 ). The enzyme also has significant stability (70%-80%) in a few organic solvents. Thus, the increased stability of the enzyme in the presence of oxidizing agent, surfactants, and organic solvents may find potential applications in the detergent industry and peptide synthesis in nonaqueous media.

Purification and characterization of a new neutral metalloprotease from marine Exiguobacterium sp. SWJS2

“…SKPB5 belonged to the cysteine family [21]. The protease from Exiguobacterium profundum BK-P23 was serine protease [24]. The neutral metalloprotease in this study might be a newprotease.…”

“…Exiguobacterium profundum BK-P23 produced a novel oxidant-and surfactant-stable extracellular alkaline protease, which was most active at pH 8.0 and 40°C [23,24]. Exiguobacterium sp.…”

SHG10 keratinolytic alkaline protease from Bacillus licheniformis SHG10 DSM 28096: Robust stability and unusual non‐cumbersome purification

“…strain AB1, B. circulans MTCC 7942, Exiguobacterium profundum BK‐P23, S. koyangensis TN 650, Caldicoprobacter guelmensis , Bacillus sp. EMB9, and B. licheniformis 3C5) . Apparently, absence of proteolytic activity in factions eluted from the column with high ionic strength of 1 M NaCl (0–1) might confirm absence of protease isozyme of different pI(s) produced by B. licheniformis SHG10 DSM 28096 under the stated conditions.…”

“…Additionally, single bind appearance of the purified enzyme in the flow through fractions conferred two conclusions. The first one states that SHG10 keratinolytic alkaline protease has one protein subunit as the majority of other proteases of microbial origins reported in the literature till now [8,9,16,24,25,[39][40][41][42]. The last conclusion underlines the absence of protease isozymes of different molecular masses.…”

“…Unlike purification folds (2.1–38.0) of other proteolytic enzymes reported in the literature, SHG10 keratinolytic alkaline protease purification fold is either relatively so higher than those of others or slightly comparable with those of others. Even though comparable purification folds of proteolytic enzymes to that of SHG10 keratinolytic alkaline protease, reported in the literature, had been obtained through a multi‐step purification strategy using at least two chromatographic columns successively apart from ammonium sulfate precipitation step . For instance, protease of B. koreensis (BK‐P21A) was purified to homogeneity through a three step purification strategy (ammonium sulfate precipitation, Superdex 200 10/300 GL, and Superdex 75 10/300 GL column chromatography) with a low fold purification of 5.0 .…”