Purification and Characterization of a New Thermostable, Haloalkaline, Solvent Stable, and Detergent Compatible Serine Protease fromGeobacillus toebiiStrain LBT 77

Abstract

A new thermostable, haloalkaline, solvent stable SDS-induced serine protease was purified and characterized from a thermophilic Geobacillus toebii LBT 77 newly isolated from a Tunisian hot spring. This study reveals the potential of the protease from Geobacillus toebii LBT 77 as an additive to detergent with spectacular proprieties described for the first time. The protease was purified to homogeneity by ammonium sulfate precipitation followed by Sephadex G-75 and DEAE-Cellulose chromatography. It was a monomeric enzyme with molecular weight of 30 kDa. The optimum pH, temperature, and NaCl for maximum protease activity were 13.0, 95°C, and 30%, respectively. Activity was stimulated by Ca2+, Mg2+, DTNB, β-mercaptoethanol, and SDS. The protease was extremely stable even at pH 13.25, 90°C, and 30% NaCl and in the presence of hydrophilic, hydrophobic solvents at high concentrations. The high compatibility with ionic, nonionic, and commercial detergents confirms the utility as an additive to cleaning products. Kinetic and thermodynamic characterization of protease revealed K m = 1 mg mL−1, V max = 217.5 U mL−1, K cat/K m = 99 mg mL−1 S−1, E a = 51.5 kJ mol−1, and ΔG ⁎ = 56.5 kJ mol−1.

Purification and partial characterization of a thermo-halostable protease produced by Geobacillus sp. strain PLS A isolated from undersea fumaroles

“…Meanwhile, a thermophilic Bacillus toebii strain LBT 77 isolated from a hot spring has optimum activity at 95°C and pH 13. The activity of the enzyme increases with the addition of 25% acetonitrile, methanol, ethanol, and n-butanol [6]. This shows the versatility of proteases for various uses.…”

“…Several previous studies report that optimum specific activities are achieved at higher ammonium sulfate concentrations, normally in the range of 50-75% [5,19]. Ammonium sulfate concentration up to 90% is sometimes needed [6].…”

“…Many studies have reported the effect of temperature on the activity of protease from thermophilic Bacillus groups. Optimum temperature ranges from as low as 50°C [5,19,24] to 95°C [6].…”

Genomic and transcriptome analyses of a thermophilic bacterium Geobacillus stearothermophilus B5 isolated from compost reveal its enzymatic basis for lignocellulose degradation

“…One unit of enzyme activity was de ned as the amount of enzyme that released 1 µmol of reducing sugars per minute. Protease activity was measured according to Thebti et al [54] with casein as substrate. One unit of protease activity was de ned as the amount of enzyme that released 1 µmol of tyrosine per minute.…”

Parageobacillus thermantarcticus, an Antarctic Cell Factory: From Crop Residue Valorization by Green Chemistry to Astrobiology Studies

“…In addition, the protease isolated by P. thermantarcticus showed stability and compatibility with some commercial detergents in the presence of CaCl 2 . The purified protease from P. thermantarcticus possesses thermophilicity, thermostability, resistance to organic solvent and detergents, with extensive activity in the pH range, characteristics that make it particularly interesting for applications in the detergent sector [82][83][84] (Table 3). Table 3.…”