[Show abstract][Hide abstract] ABSTRACT: An optimum Aspergillus oryzae CBS 819.72 α-amylase production in a laboratory-scale fermentor using a wheat grinding by-product as a sole carbon source (340 U/mL) was obtained after 48 h of batch fermentation under an agitation rate of 900 rpm and a pH maintained constant for 24 h. The application of this α-amylase preparation at an adequate concentration showed positive effects on dough properties and bread quality. Extensographic analysis revealed that while this addition induced a significant increase in maximal dough resistance to extension and area below the curve (energy), it brought a substantial decrease in extensibility. Farinographic results revealed small decreases in terms of time development, water absorption, and dough stability. Bread volume was also observed to undergo a significant increase.
[Show abstract][Hide abstract] ABSTRACT: An extracellular keratinolytic protease (SAPDZ) was produced and purified from a newly isolated Bacillus
circulans strain DZ100. Matrix assisted laser desorption ionization-time of flight mass spectrometry
(MALDI-TOF/MS) analysis revealed that the purified enzyme is a monomer with a molecular mass of
32019.10 Da. The sequence of the 25 N-terminal residues of SAPDZ showed high homology with those of
Bacillus proteases. Optimal activity was achieved at pH 12.5 and 85 �C. This enzyme was completely
inhibited by phenylmethanesulfonyl fluoride (PMSF) and diiodopropyl fluorophosphates (DFP), which
suggests that it belongs to the serine protease family. Compared to the other tested proteases, SAPDZ
exhibited broader substrate specificity, higher levels of catalytic efficiency, and greater keratinolytic
activity, which made it able to accomplish the entire feather-biodegradation process on its own. The
sapDZ gene encoding SAPDZ was cloned, sequenced, and expressed in Escherichia coli. The biochemical
properties exhibited by the extracellular purified recombinant enzyme (rSAPDZ) were similar to those of
the native one. Above all, SAPDZ exhibited marked stability to detergents, making it a potential candidate
for future applications in detergent formulations and an efficient eco-friendly enzyme for the biodegradation
of feather keratin.
[Show abstract][Hide abstract] ABSTRACT: An extracellular alkaline elastase was produced from Pseudomonas aeruginosa CTM50182. It was chromatographically purified using HPLC and Mono Q Sepharose column. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis revealed that the purified enzyme (called AMPP) was a monomer with a molecular mass of 33015.18Da. The N-terminal 29 amino acid sequence of AMPP showed high homology with those of Pseudomonas elastases. It showed optimal activity at pH 12 and 80°C and was stable at a pH range of 9 to 12 after 120h of incubation. Its thermoactivity and thermostability were upgraded in the presence of 5mM Co(2+). Its half-life times at 70 and 80°C were 16 and 10h, respectively. It was completely inhibited by ethylene glycol-bis (β-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), and 1,10-phenanthroline, suggesting that it belongs to the metalloprotease family. AMPP also exhibited high catalytic efficiency, organic solvent-tolerance, and hydrolysis. The lasB gene encoding AMPP was cloned, sequenced, and expressed in Escherichia coli. The biochemical properties of the extracellular purified recombinant enzyme (rAMPP) were similar to those of native AMPP. This organic solvent-stable protease could be considered a potential candidate for application as a biocatalyst in the synthesis of enzymatic peptides.
International journal of biological macromolecules 05/2013; 60. DOI:10.1016/j.ijbiomac.2013.05.019 · 2.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The partition and purification of α-amylase from a culture supernatant of Aspergillus oryzae CBS 819.72 was made in aqueous two-phase system (ATPS). According to bibliography and preliminary studies, the factors polyethylene glycol (PEG) molecular weight (MPEG) and concentration (CPEG), buffer type (BU) and concentration (CBU), temperature (T), salt nature (SALT) and concentration (CSALT), bioligand (BL) and concentration (CBL) and pH were investigated using a Plackett–Burman design to identify the factors affecting separation. Taking into consideration a simultaneous increase in enzyme recovery (RY) and purification factor (PF), the best performance of the system was obtained at 4 °C and pH 6 using PEG 8000 g/mol, citrate buffer, KCl and sucrose. Experimental Box–Behnken design together with the Response Surface Methodology (RSM) have been used to find optimum CPEG, CCitrate and CSALT. Quadratic models were predicted for PF and RY in the top phase and a better compromise between these two parameters can be found by superimposing the contour plots of PF and RY for 8% citrate. A region in the experimental space can be defined where the purification factor is always higher than 3 with yields exceeding 65%.
[Show abstract][Hide abstract] ABSTRACT: The newly Tunisian soil-isolated bacterium, producing the alkaline proteinase termed SAPB that was already purified and characterized
, was assigned as Bacillus pumilus CBS strain on the basis of biochemical properties and 16S rRNA gene sequencing. The maximum protease activity recorded after
24 h of incubation in an optimized medium at 37°C was 6,500 U/mL in shaking flask culture and 25,000 U/mL in fermentor. SAPB
showed excellent stability and compatibility in laundry detergent retaining more than 98% of its initial activity after pre-incubation
for 1 h at 40°C with Det, followed by OMO (97%), Dinol (94%), and Dixan (93%). Examination of various stained cloth pieces
exhibited a remarkable efficiency in the removal of blood and chocolate stains. More interestingly, SAPB demonstrated powerful
dehairing capabilities of hair removal from skin with minimal damage on the collagen and a nearly complete feather-degradation.
Likewise, Bacillus pumilus CBS effectively degraded feather-meal (98.5%), chicken feather (92%), goat hair (80%), and bovine hair (68%) whereas sheep
wool under went less degradation. Keratin-degradation resulted in sulfhdryl group formation (0.95∼3.91 μM).
-Bacillus pumilus CBS
[Show abstract][Hide abstract] ABSTRACT: The production optimization of alpha-amylase (E.C.188.8.131.52) from Aspergillus oryzae CBS 819.72 fungus, using a by-product of wheat grinding (gruel) as sole carbon source, was performed with statistical methodology based on three experimental designs. The optimisation of temperature, agitation and inoculum size was attempted using a Box-Behnken design under the response surface methodology. The screening of nineteen nutrients for their influence on alpha-amylase production was achieved using a Plackett-Burman design. KH(2)PO(4), urea, glycerol, (NH(4))(2)SO(4), CoCl(2), casein hydrolysate, soybean meal hydrolysate, MgSO(4) were selected based on their positive influence on enzyme formation. The optimized nutrients concentration was obtained using a Taguchi experimental design and the analysis of the data predicts a theoretical increase in the alpha-amylase expression of 73.2% (from 40.1 to 151.1 U/ml). These conditions were validated experimentally and revealed an enhanced alpha-amylase yield of 72.7%.
[Show abstract][Hide abstract] ABSTRACT: A bacterial strain designed US132, isolated from a Tunisian soil was selected for its production of a potent cyclodextrin glycosyltransferase (CGTase) activity. This strain was identified as Paenibacillus pabuli by sequencing of the 16S rDNA and the 16S-23S internal transcribed spacer (ITS). The US132 CGTase, purified to homogeneity by hydrophobic interaction chromatography and starch adsorption, is a monomer of approximately 70 kDa. This enzyme exhibited a maximal activity at 65 °C, in presence of 10 mM calcium, and was most active at pH range 5.5–9 with an optimum at 6.5. Using 10% (w/v) of potato starch, this CGTase produced a high level of cyclodextrins reaching 42 g/l with a β-cyclodextrin ratio of 63%. Furthermore, this enzyme can be used in the bread-baking process since its addition in the dough mix improved significantly the loaf volume and decreased the firmness of bread during storage.
[Show abstract][Hide abstract] ABSTRACT: A new aerobic Gram-positive bacterium designated TN58 producing antibacterial activities against Gram-positive and Gram-negative bacteria was isolated from Tunisian soil. The nucleotide sequence of the 16S rRNA gene (1516 bp) of the TN58 strain showed high similarity (96-98%) to the Streptomyces 16S rRNA genes, especially with that of Streptomyces lavendulae which produces the anti-tumor compound mitomycin C, and the cyclic peptide antibiotic, complestatin. Cultural characteristic studies, alignment data of the 16S rRNA gene, and analysis of the nucleotide sequence of a 2.2 kb genomic DNA fragment from TN58 strongly suggested that this strain could be an actinomycete and most probably belongs to the genus Streptomyces. Study of the influence of different nutritional compounds on antibiotic production showed that the highest antibacterial activities were obtained when glycerol at 1% (w/v) was used as sole carbon source in the presence of potassium. In analytical conditions, the application to supernatant culture of the TN58 strain of various extraction and purification steps led to the isolation of two pure active molecules having a retention time of 38.6 and 50.2 min, respectively. TN58 strain was untransformable with the Streptomyces cloning vector pIJ702 via classical polyethylene glycol (PEG) protoplast transformation and previously described Streptomyces electroporation procedures. Transformation was rendered possible by the electroporation technique only after utilization of a preculture medium without sucrose and a regeneration plate containing a low sucrose concentration.
Current Microbiology 01/2005; 49(6):400-6. DOI:10.1007/s00284-004-4292-9 · 1.42 Impact Factor