Cleaner degreasing of sheepskins by the Yarrowia lipolytica LIP2 lipase as a chemical-free alternative in the leather industry.

Conventional degreasing of skins and hides in the leather industry requires high amounts of organic solvents and detergents that cause environmental issues. In this study, the LIP2 lipase from the yeast Yarrowia lipolytica (YLLIP2) was shown to be effective in degreasing sheepskins, thus reducing the amount of harmful chemicals. Using 6 mg of lipase/kg of raw skin, successful degreasing was achieved in only 15 min at pH 8 and 30°C. ToF-SIMS mass spectra of chemically and enzymatically treated sheepskins are consistent with a selective elimination process for the enzymatic treatment. Comparative SEM microscopy, ATR-FTIR spectroscopy and physicochemical analyses showed better properties of the enzymatically treated leather than those of the chemical treatment. Effluent physicochemical parameters showed that the enzymatic treatment is a cleaner degreasing operation. Altogether, this work opens new horizons to use the YLLIP2 lipase as a more efficient alternative in the leather industry.

Production, purification, and biochemical characterization of serine alkaline protease from Penicillium chrysogenium strain X5 used as excellent bio-additive for textile processing.

A new ascomycete fungus X5, a hyperproducer (9000 U/mL) of a serine alkaline protease (SAPTEX) was identified as Penicillium chrysogenum. The experimental purification protocol comprises three steps: heat treatment (10 min at 80 °C) followed by an ammonium sulfate precipitation (30-50%)-dialysis, and a UNO Q-12 anion exchange chromatography using the FPLC system. The chemical characterizations performed include physico-chemical determination and spectroscopic analysis. The MALDI-TOF/MS analysis revealed that the purified enzyme was a monomer with a molecular mass of 43,074.11 Da. The 25 residue NH2-terminal sequence of the enzyme showed high homology with Penicillium proteases. The optimum pH and temperature values for protease activity were pH 10 and 80 °C, respectively. Compared to other proteases (SPTC; Flavourzyme® 500 L; Proteinase, type XXIII; Proteinase K; and Alcalase® 2.4 L), SAPTEX has the highest catalytic efficacy, hydrolysis degree, and a powerful stability toward some commercial detergents. According to morphological, physico-chemical [scanning electron microscopy (SEM), energy dispersive X-Ray analysis (EDX), and FTIR-Fourier transform infrared spectroscopy], and mechanical evaluation, SAPTEX has no destructive impact on fibers after the enzyme treatment and a very slight effect on textile support. Obtained results suggested that SAPTEX may be considered as a potential candidate as a protein stain removal product for textile supports.

Characterization of a novel protease from Aeribacillus pallidus strain VP3 with potential biotechnological interest.

The present study investigates the purification and physico-chemical characterization of an extracellular protease from the Aeribacillus pallidus strain VP3 previously isolated from a geothermal oil-field (Sfax, Tunisia). The maximum protease activity recorded after 22h of incubation at 45°C was 3000U/ml. Pure enzyme, designated as SPVP, was obtained after ammonium sulfate fractionation (40-60%)-dialysis followed by heat-treatment (70°C for 30min) and UNO Q-6 FPLC anion-exchange chromatography. The purified enzyme is a monomer of molecular mass about 29kDa. The sequence of the 25 NH2-terminal residues of SPVP showed a high homology with those of Bacillus proteases. The almost complete inhibition by PMSF and DIFP confirmed that SPVP is a member of serine protease family. Its optima of pH and temperature were pH 10 and 60°C, respectively. Its half-life times at 70 and 80°C were 8 and 4h, respectively. Its catalytic efficiency was higher than those of SAPCG, Alcalase Ultra 2.5L, and Thermolysin type X. SPVP exhibited excellent stability to detergents and wash performance analysis revealed that it could remove blood-stains effectively and high resistance against organic solvents. These properties make SPVP a potential candidate for applications in detergent formulations and non-aqueous peptide biocatalysis.

Biochemical and molecular characterization of new keratinoytic protease from Actinomadura viridilutea DZ50.

A new extracellular thermostable keratinolytic protease, designated KERDZ, was purified and characterized from a thermophilic actinomycetes Actinomadura viridilutea DZ50 isolated from Algerian fishing port. The isolate exhibited high keratinase production when grown in chicken-feather meal media (18,000U/ml) after 96-h of incubation at 45°C. The enzyme was purified by ammonium sulfate precipitation (35-55%)-dialysis and heat treatment (30min at 75°C) followed by UNO S-1 FPLC cation exchange chromatography and size exclusion HPLC column. The biochemical characterizations carried on include physico-chemical determination and spectroscopic analysis. The MALDI-TOF/MS analysis revealed that the purified enzyme was a monomer with a molecular mass of 19536.10-Da. The sequence of the 25 N-terminal residues of KERDZ showed high homology with those of actinomycetes keratinases. Optimal activity was achieved at pH 11 and 80°C. KERDZ was completely inhibited by PMSF and DFP suggested its belonging to the serine keratinase family. KERDZ displayed higher levels of hydrolysis and catalytic efficiency than bacterial keratinases (KERAK-29, Actinase E, and KERAB) and subtilisins (subtilisin Carlsberg and subtilisin Novo). The kerDZ gene encoding KERDZ was isolated and its DNA sequence was determined. These properties make KERDZ a potential, promising and eco-friendly alternative to the conventional chemicals used for industrial applications.

A novel organic solvent- and detergent-stable serine alkaline protease from Trametes cingulata strain CTM10101.

A protease-producing fungus was isolated from an alkaline wastewater of chemical industries and identified as Trametes cingulata strain CTM10101 on the basis of the ITS rDNA gene-sequencing. It was observed that the fungus strongly produce extracellular protease grown at 30°C in potato-dextrose-broth (PDB) optimized media (13500U/ml). The pure serine protease isolated by Trametes cingulata (designated SPTC) was purified by ammonium sulfate precipitation-dialysis followed by heat-treatment and UNO S-1 FPLC cation-exchange chromatography. The chemical characterization carried on include phisico-chemical determination and spectroscopie analysis. The MALDI-TOF/MS analysis revealed that the purified enzyme was a monomer with a molecular mass of 31405.16-Da. The enzyme had an NH2-terminal sequence of ALTTQTEAPWALGTVSHKGQAST, thus sharing high homology with those of fungal-proteases. The optimum pH and temperature values of its proteolytic activity were pH 9 and 60°C, respectively, and its half-life times at 60 and 70°C were 9 and 5-h, respectively. It was completely inhibited by PMSF and DFP, which strongly suggested its belonging to the serine protease family. Compared to Flavourzyme(®)500L from Aspergillus oryzae and Thermolysin typeX from Geobacillus stearothermophilus, SPTC displayed higher levels of hydrolysis, substrate specificity, and catalytic efficiency as well as elevated organic solvent tolerance and considerable detergent stability. Finally, SPTC could potentially be used in peptide synthesis and detergent formulations.