共查询到20条相似文献,搜索用时 15 毫秒
1.
Richard A. Dilley Gerald A. Peters Elwood R. Shaw 《The Journal of membrane biology》1972,8(1):163-180
Summary Spinach chloroplasts were exposed to35S-labeledp-(diazonium)-benzenesulfonic acid (DABS), a water soluble compound which does not penetrate lipophilie regions of membranes, and which is highly reactive toward amino acid functionagroups such as -amino, sulfhydryl, histidine, and tyrosine groups. Amino groups inl lipids can also form similar, stable covalent bonds by diazo coupling. Both chloroplast lipids and proteins were labeled with DABS, the total binding being about 1 DABS per 10 chlorophylls, depending on the reaction conditions.After diazo coupling and subsequent digitonin fractionation into photosystems I and II enriched fractions, it was observed that PS-I was more highly labeled than PS-III usually by a factor of 10 to 24 times (on a per chlorophyll basis). After digitonin isolation, however, the PS-II portion bound an amount of DABS similar to the PS-I binding, We interpret these data as consistent with the binary membrane hypothesis (Arntzen. Dilley and Crane (1969),J. Cell Biol.
43:16), which visualizes PS-I on the externa, half of a 90 Å grana membrane, and PS-II occurring on the interior half of thel membrane. The alternative explanation that PS-II and PS-I are arranged as a mosaic, and that the low DABS binding in PS-II is caused by burial of the diazo reactive groups in the interior of the proteins (and only exposed through the denaturing effect of digitonin) is not directly ruled out. However, this alternative is not consistent with the facts that: (a) most of the membrane proteins in PS-I and PS-II are identical in electrophoretic properties and therefore probably have similar overall structures; and (b) digitonin does not lead to appreciable denaturation of proteins, evidenced by the retention of PS-II electron transport activity. 相似文献
2.
Werner Hummel Horst Schütte Maria-Regina Kula 《Applied microbiology and biotechnology》1981,12(1):22-27
Summary To develop a large-scale isolation of leucine dehydrogenase (E.C. 1.4.1.9) as industrial catalyst we carried out a limited screening for microorganisms with high leucine dehydrogenase activity.Conditions for the growth and enzyme formation of Bacillus sphaericus (DSM 396) which proved to be the best enzyme producer were optimized. The highest yield in volume and specific activity were obtained using glucose and yeast-extract in the medium.The highest specific enzyme activity was found at the end of the exponential growth phase. Cultivation of Bacillus sphaericus under optimal conditions increased the yield to about 3 U mg–1. The heat stability of the enzyme was exploited to develop a simple large-scale purification. Together with an ultrafiltration step, the enzyme could be enriched 9fold in a short time. After further purification using DE-cellulose an enzyme preparation (25fold enriched) was obtained; suitable as a technical catalyst in amino acid production. 相似文献
3.
The optimization of operating parameters for the isolation of peroxidase from horseradish (Armoracia rusticana) roots with ultrafiltration (UF) technology was systemically studied. The effects of UF operating conditions on the transmission of proteins were quantified using the parameter scanning UF. These conditions included solution pH, ionic strength, stirring speed and permeate flux. Under optimized conditions, the purity of horseradish peroxidase (HRP) obtained was greater than 84 % after a two-stage UF process and the recovery of HRP from the feedstock was close to 90 %. The resulting peroxidase product was then analysed by isoelectric focusing, SDS–PAGE and circular dichroism, to confirm its isoelectric point, molecular weight and molecular secondary structure. The effects of calcium ion on HRP specific activities were also experimentally determined. 相似文献
4.
Byeong Jo Min Yang Soon Park Seong Woo Kang Yoon Seok Song Jong Ho Lee Chulhwan Park Chan Wha Kim Seung Wook Kim 《Biotechnology and Bioprocess Engineering》2007,12(3):302-307
Medium composition was optimized for the production of xylanase byAspergillus niger KK2 using statistical experimental designs. Corn steep liquor (CSL) and industrial yeast extract (IYE) were the most important
factors affecting xylanase activity. The medium that produced the optimum conditions for the production of xylanase contained
3% rice straw, 1% wheat bran, 6.3% CSL, 0.15% IYE, and 0.5% KH2PO4. After 4 days of cultivation under optimized conditions in a 2.5-L stirred tank reactor the activity and productivity of
xylanase were 620 IU/mL and 6,458 IU/L.h, respectively. The highest xylanase activity obtained using the optimized medium
was 80% greater than the activity obtained using basal medium. The xylanase activity predicted by a polynomial model was 670
IU/ml. 相似文献
5.
The optimization of operating conditions for cellulose hydrolysis was systemically undertaken using an ultra-scaled down membrane
bioreactor based on the parameter scanning ultrafiltration apparatus. The bioconversion of cellulose saccharification was
carried out with freely suspended cellulase from Aspergillus niger as the biocatalyst. The polyethersulfone ultrafiltration membranes with a molecular weight cutoff of 10 kDa were used to
construct the enzymatic membrane bioreactor, with the membrane showing a complete retaining of cellulase and cellobiase. The
influence of solution pH, temperature, salt (NaCl) concentration, presence of cellobiase, cellulose-to-enzyme ratio and stirring
speed on reducing sugar production was examined. The results showed that the addition of an appropriate amount of NaCl or
cellobiase had a positive effect on reducing sugar formation. Under the identified optimal conditions, cellulose hydrolysis
in the enzymatic membrane bioreactor was tested for a long period of time up to 75 h, and both enzymes and operation conditions
demonstrated good stability. Also, the activation energy (E
a) of the enzymatic hydrolysis, with a value of 34.11 ± 1.03 kJ mol−1, was estimated in this study. The operational and physicochemical conditions identified can help guide the design and operation
of enzymatic membrane bioreactors at the industrial scale for cellulose hydrolysis. 相似文献
6.
Optimization of bioethanol production during simultaneous saccharification and fermentation in very high-gravity cassava mash 总被引:1,自引:0,他引:1
Hydrolysis and fermentation conditions for production of ethanol from very high-gravity cassava mash by Saccharomyces cerevisiae during simultaneous saccharification and fermentation (SSF) processing were optimized using a statistical methodology. During
the first part of the study, Placket–Burman design (PBD) was used to study 19 factors that could potentially influence ethanol
production. Gravity, particle size, initial pH, and fermentation temperature were identified as key factors that significantly
increased final ethanol concentration. The main and interaction effects of these factors were subsequently evaluated based
on a quadratic equation generated by central composite design (CCD) using response-surface methodology (RSM). Under the optimized
very high-gravity conditions, the final ethanol concentration obtained from experiment increased from 8.21% (wt.%) to 15.03%
(wt.%) and was in good agreement with model prediction. By employing two other commercial Saccharomyces strains, similar results were obtained under the same optimized condition. Therefore, we conclude that final ethanol concentration,
ethanol productivity (V
P/max), glucose utilization (Y
G/s, Y
P/s), and fermentation efficiency (η
f) were enhanced or maintained under the optimized condition of 40% gravity, 390 μm particle size, initial pH 5.5, and 27°C
fermentation temperature. 相似文献
7.
Cassava (Manihot esculenta Crantz) is a known source of linamarin, but difficulties associated with its isolation have prevented it from being exploited as
a major source. A batch adsorption process using activated carbon proved successful in its isolation, with ultrafiltration
playing a pivotal role in its purification. Thirty-two minutes of contact time was required for 60 g of extract, yielding
1.7 g of purified product. Picrate paper, infra-red and 1HNMR analysis confirmed the presence and structure of linamarin. Cytotoxic effects of linamarin on MCF-7, HT-29 and HL-60
cells were determined using the MTT assay. Cytotoxic effects were significantly increased in the presence of linamarase (β-glucosidase),
with a 10–fold decrease in the IC50 values obtained for HL-60 cells. This study thus describes a method for the isolation and purification of linamarin from
cassava, as well as its cytotoxicity potential. 相似文献
8.
When grown heterotrophically in the dark on enriched culture medium, the pigment-deficient strain of Scenedesmus obliquus, mutant C-6E, is uniquely characterized by a complete deficiency in carotenoids and chlorophyll b while retaining a low level of chlorophyll a which is exclusively utilized in photosystem I-type reactions. The strain lacks photosystem II activity but exhibits all PS-I reactions tested, including P700 redox reactions, photoreduction of CO2 with hydrogen as electron donor, and O2 uptake following methyl viologen reduction. The mutant contains 10 times more P700 per chlorophyll than the wild type and develops the pigment-protein complex of PS-I, CP-I. The action spectrum for methyl viologen reduction compares favorable to the low temperature absorption spectrum of whole cells. Both the chlorophyll fluorescence excitation and emission spectra of pigment-protein complexes derived from cells of C-6E show patterns typical of PS-I. The strain lacks the LHCs and CP-II as well as their respective apoproteins. The absence of carotenoids appears to prevent the development of the normal variety of pigment-protein complexes and the accumulation of Chl b. This inability is also expressed by the presence of only single stranded thylakoid membranes in the chloroplast of C-6E. When heterotrophically grown cells of this mutant are exposed to white light of 8 or 22 W m?2, 50% of its chlorophyll is lost by photooxidation within 4 or 1.5 hours, respectively. 相似文献
9.
Fabiano Jares Contesini Vania Castriani Fernades da Silva Rafael Ferreira Maciel Rosemary Joana de Lima Francisco Fábio Cavalcante Barros Patrícia de Oliveira Carvalho 《Journal of microbiology (Seoul, Korea)》2009,47(5):563-571
The lipase produced by the Aspergillus niger strain AC-54 has been widely studied due to its enantioselectivity for racemic mixtures. This study aimed to optimize the
production of this enzyme using statistical methodology. Initially a Plackett-Burman (PB) design was used to evaluate the
effects of the culture medium components and the culture conditions. Twelve factors were screened: water content, glucose,
yeast extract, peptone, olive oil, temperature, NaH2P04, KH2P04, MgS04-7H20, CaCl2, NaCI, and MnS04. The screening showed that the significant factors were water content, glucose, yeast extract, peptone, NaH2P04, and KH2P04, which were optimized using response surface methodology (RSM) and a mathematical model obtained to explain the behavioral
process. The best lipase activity was attained using the following conditions: water content (20%), glucose (4.8%), yeast
extract (4.0%), and NaH2P04 (4.0%). The predicted lipase activity was 33.03 U/ml and the experimental data confirmed the validity
of the model. The enzymatic activity was expressed as μmoles of oleic acid released per minute of reaction (μmol/min). 相似文献
10.
Optimal operating policy of the ultrafiltration membrane bioreactor for enzymatic hydrolysis of cellulose 总被引:1,自引:0,他引:1
The dilution rate of an ultrafiltration membrane bioreactor in the enzymatic hydrolysis of cellulose was optimized using the kinetic model developed by Fan and Lee.(4) The sequence of optimal dilution rates was found to generally consist of an initial period of a minimal value (batch period), a subsequent period of maximum dilution rate, a period of a second batch, and a final period of a singular dilution rate. The effects of operating conditions, such as beta-glucosidase activity, operating time, maximum dilution rate, substrate feeding rate, and enzyme-to-substrate ratio on both the conversion yield and the sequence of optimal dilution rates were investigated. To evaluate the validity of kinetic model employed in this work, enzymatic hydrolysis was carried out using alpha-cellulose as a substrate in the ultrafiltration membrane bioreactor. The experimental data were well consistent with the simulation results. (c) 1993 John Wiley & Sons, Inc. 相似文献
11.
Optimization of Nutrient and Fermentation Parameters for Antifungal Activity by Streptomyces lavendulae Xjy and Its Biocontrol Efficacies against Fulvia fulva and Botryosphaeria dothidea
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This study has been proved that statistical experimental designs offer efficient and feasible approaches for determination of culture medium and fermentation conditions of Streptomyces lavendulae Xjy. Nutrition and cultural conditions have high influence on the antibiotic production of S. lavendulae Xjy. Soya bean meal, NaCl and (NH4)2SO4 were identified as the best ingredients for high antifungal activity of Xjy based on single variable experiments. Using response surface methodology, the best concentrations of medium ingredients were 1.46% soya bean meal, 2.25% NaCl and 1.26% (NH4)2SO4. The potential components were reduced from seven to three, and the antibiotic activity was increased by 10%. The optimized conditions for Xjy in a 250‐ml Erlenmeyer flask were initial pH 5, medium volume 25 ml/250 ml and inoculation volume 10% at 30°C with 200 rpm by response surface methodology. Validation experiments carried out to check the accuracy of the models indicated that the predicted values agreed with the experimental values. The extension of necrosis lesion of Botryosphaeria dothidea was impeded by the application of Xjy in apple fruit. In tomato plants, Xjy showed the 43.7–51.7% curative and protective effects against Fulvia fulva. The optimal medium not only gave a 20% increase in antifungal activity, but it provided a simpler formulation. The results will set a basis for further study on large‐scale fermentation volumes using S. lavendulae Xjy and be useful for the development of more advanced control strategies on plant diseases. 相似文献
12.
Xiao-Jun Huang An-Guo Yu Jun Jiang Chao Pan Jing-Wen Qian Zhi-Kang Xu 《Journal of Molecular Catalysis .B, Enzymatic》2009,57(1-4):250-256
In this work, poly(acrylonitrile-co-acrylic acid) (PANCAA) was electrospun into nanofibers with a mean diameter of 180 nm. To create a biofriendly microenvironment for enzyme immobilization, collagen or protein hydrolysate from egg skin (ES) was respectively tethered on the prepared nanofibrous membranes in the presence of 1-ethyl-3-(dimethyl-aminopropyl) carbodiamine (EDC)/N-hydroxyl succinimide (NHS). Confocal laser scanning microscopy (CLSM) was used to verify the surface modification and protein density on the nanofibrous membranes. Lipase from Candida rugosa was then immobilized on the protein-modified nanofibrous membranes by covalent binding using glutaraldehyde (GA) as coupling agent, and on the nascent PANCAA nanofibrous membrane using EDC/NHS as coupling agent, respectively. The properties of the immobilized enzyme were assayed. It was found that different pre-tethered biomacromolecules had distinct effects on the immobilized enzyme. The activity retention of the immobilized lipase on ES hydrolysate-modified nanofibrous membrane increased from 15.0% to 20.4% compared with that on the nascent one, while it was enhanced up to more than quadrupled (activity retention of 61.7%) on the collagen-modified nanofibrous membrane. The kinetic parameter, Km and Vmax, were also determined for the free and immobilized lipases. Furthermore, the stabilities of the immobilized lipases were obviously improved compared with the free one. 相似文献
13.
Chantal Brämer Kimia Ekramzadeh Frank Lammers Thomas Scheper Sascha Beutel 《Biotechnology progress》2019,35(4):e2812
The natural production of patchouli oil in developing countries cannot meet the increasing demand any more. This leads to socioecological consequences, such as the use of arable land, which is actually intended for food. Hence, the world market price increased up to $150/kg. An alternative is the biotechnological production of patchouli oil using a multiproduct sesquiterpene synthase, the patchoulol synthase (PTS). Here, we report the optimization of recombinant PTS purification from Escherichia coli lysate using continuous immobilized metal affinity chromatography. First, the purification conditions of the batch process were optimized in regard to optimal buffer composition and optimized chromatographic conditions. The best purification result was achieved with Co2+-immobilized metal affinity chromatography (Sartobind® IDA 75) with a triethanolamine buffer at pH 7, 0.5 M NaCl, 10% [vol/vol] glycerol, 5 mM MgCl2 and 250 mM imidazole for product elution. This optimized method was then transferred to a continuous chromatography system using three membrane adsorber units (surface of 75 cm2 each). Within 1.5 hr in total, 4.55 mg PTS with a final purity of 98% and recovery of 68% could be gained. The purified enzyme was used to produce 126 mg/L (-)-patchoulol from farnesyl pyrophosphate. Here, for the first time bioactive PTS was successfully purified using membrane adsorbers in a continuous downstream process. 相似文献
14.
Stability of membrane protein is crucial during protein purification and crystallization as well as in the fabrication of protein-based devices. Several recent studies have examined how various surfactants can stabilize membrane proteins out of their native membrane environment. However, there is still no single surfactant that can be universally employed for all membrane proteins. Because of the lack of knowledge on the interaction between surfactants and membrane proteins, the choice of a surfactant for a specific membrane protein remains purely empirical. Here we report that a group of short amphiphilic peptides improve the thermal stability of the multi-domain protein complex photosystem-I (PS-I) in aqueous solution and that the peptide surfactants have obvious advantages over other commonly used alkyl chain based surfactants. Of all the short peptides studied, Ac-I5K2-CONH2 (I5K2) showed the best stabilizing effect by enhancing the melting temperature of PS-I from 48.0°C to 53.0°C at concentration of 0.65 mM and extending the half life of isolated PS-I significantly. AFM experiments showed that PS-I/I5K2/Triton X-100 formed large and stable vesicles and thus provide interfacial environment mimicking that of native membranes, which may partly explain why I5K2 enhanced the thermal stability of PS-I. Hydrophobic and hydrophilic group length of IxKy had an important influence on the stabilization of PS-I. Our results showed that longer hydrophobic group was more effective in stabilizing PS-I. These simple short peptides therefore exhibit significant potential for applications in membrane protein studies. 相似文献
15.
Muhamad O. Al-Limoun Khalid Y. Alsharafa Haitham N. Qaralleh Suddam A. Alrawashdeh 《Biocatalysis and Biotransformation》2019,37(2):139-151
The extracellular lipase produced by Acinetobacter sp. K5b4 was purified to homogeneity using ultrafiltration (cutoff 30?KDa) followed by gel filtration chromatography on Sephadex G-50. The enzyme was purified to homogeneity with an apparent molecular mass of 133?KDa by SDS-PAGE. This purification resulted on 10.24 fold with 18.3% recovery. The Km and Vmax of purified enzyme when using pNPL hydrolysis were 4.0?mM and 73.53?nmol/ml/min, respectively. The pure enzyme was greatly stimulated in the presence of 20, 40 and 60% (v/v) methanol, DMSO and acetone whereas, ethanol, acetonitrile and propanol decreased the enzyme activity. Maximum enzyme activity was achieved at pH 7.0 and incubation temperature of 27?°C. The enzyme was stable within a pH range of 6.5 to 7 at 27?°C for 1?h. The enzyme activity was enhanced up to 36% by KCl, BaCl2, MgCl2 and CaCl2 while obviously inhibited (10–20%) by CoCl2, ZnCl2, MnCl2 and CuCl2. No inhibitory effects were observed with 1.0 and 5.0?mM of 2-mercaptoethanol and EDTA. Similarly, SDS at 1.0?mM does not affect the enzyme activity while high reduction (80%) was observed at 5.0?mM SDS concentration. The enzyme was active against p-nitrophenyl esters of C8, C12 and C16 with highest preference to the medium carbon chain p-nitrophenyl caprylate (C8). The fact that the enzyme displays distinct stability in the presence of methanol, DMSO and acetone suggests that this lipase is suitable as biocatalyst in organic synthesis where such hydrophilic organic solvents are used as a reaction media. 相似文献
16.
N I Shutilova I G Kadoshnikova N G Kozlovskaia A V Klevanik D A Zakrzhevski? 《Biokhimii?a (Moscow, Russia)》1979,44(7):1160-1171
The maximal total release of pigment protein-lipid complexes (PPLC) during their isolation from pea chloroplasts was achieved by 1-hr solubilization with Triton X-100, the Triton:chlorophyll (T/Chl) ratio being 50 mg/mg/ml. The total yield of the reaction center complexes (sigma PPLC RC) was 22,3%, whereas that of the auxiliary light-accumulating complex (ALA-PPLC) was approximately 32% with respect to Chl. An increase in the solubilization time and of the T/Chl ratio resulted in dissociation of ALA-PPLC. On the contrary, the reaction center complexes steadily maintained their composition and high photochemical activity within a wide range of T/Chl during 24--28 hrs of solubilization. The purest preparations of PPLC RC of phostosystem I (PS-I) were obtained by 24 hr-incubation (T/Chl = 80); their Chl/P700 ratio after a single fractionation on DEAE-cellulose was equal to 36. A considerable increase of T/Chl and of the solubilization time hampered the chromatographical separation of PPLC RC of PS-I and PPLC RC of PS-II. The optimal conditions for isolation of PPLC RC of PS-I and PPLC RC of PS-II were: solubilization at T/Chl 80--120 and prolongation of incubation time from 5 to 7 hrs. The photochemical activity of the complexes obtained was maximal and correlated with the minimal content of admixture P700 (1 molecule of P700 per 450--500 molecules of Chl.). 相似文献
17.
H. Li W. Q. Liang Z. Y. Wang N. Luo X. Y. Wu J. M. Hu J. Q. Lu X. Y. Zhang P. C. Wu Y. H. Liu 《World journal of microbiology & biotechnology》2006,22(1):1-7
Summary Response surface methodology was applied to optimize medium components for maximum production of a thermostable α-galactosidase by thermotolerant Absidia sp. WL511. First, the Plackett-Burman screening design was used to evaluate the effects of variables on enzyme production. Among
these variables, MgSO4 and soybean meal were identified as having the significant effects (with confidence level (90%). Subsequently, the concentrations
of MgSO4 and soybean meal were further optimized using central composite designs. The optimal parameters were determined by response
surface and numerical analyses as 0.0503% (g/g) MgSO4 and 0.406% (g/g) soybean meal and allowed α-galactosidase production to be increased from 4.4 IU g−1 to 117.8 IU g−1. The subsequent verification experiments confirmed the validity of the model. The optimum pH of enzymatic activity was 7.5
and the enzyme was stable at pH values ranging from 5.0 to 9.0. The optimum temperature was 73 °С. The enzyme was fairly stable
at temperatures up to 60 °С and had 87% of its full activity at 65 °С after 2 h of incubation. 相似文献
18.
Shripathi Bhat Alok Patel Paramananda Das Prem K. Meher Bidu R. Pillai P. Jayasankar 《Conservation Genetics》2009,10(5):1473-1475
This paper reports on isolation and characterization of 8 microsatellite loci from a partial genomic library of Macrobrachium rosenbergii using DIG labeled dinucleotides, (GT)15 and (CT)15 as probes. Primers were designed and PCR conditions optimized for repeat arrays. Polymorphism was studied using 24 individuals
collected from the wild. All the loci were polymorphic with number of alleles ranging from 3 to 5 and observed heterozygosity
0.50–0.85. All the loci except one were in agreement with Hardy–Weinberg equilibrium. No significant pair wise linkage disequilibrium
was found among the loci. These markers should prove useful for characterization of natural population as well as brood stock
management of this species. 相似文献
19.
Ricardo Sposina Sobral Teixeira Félix Gonçalves Siqueira Marcelo Valle de Souza Edivaldo Ximenes Ferreira Filho Elba Pinto da Silva Bon 《Journal of industrial microbiology & biotechnology》2010,37(10):1041-1051
This study presents data on the production, purification, and properties of a thermostable β-xylanase produced by an Aspergillus awamori 2B.361 U2/1 submerged culture using wheat bran as carbon source. Fractionation of the culture filtrate by membrane ultrafiltration
followed by Sephacryl S-200 and Q-Sepharose chromatography allowed for the isolation of a homogeneous xylanase (PXII-1), which
was 32.87 kDa according to MS analysis. The enzyme-specific activity towards soluble oat spelt xylan, which was found to be
490 IU/mg under optimum reaction conditions (50°C and pH 5.0–5.5), was 17-fold higher than that measured in the culture supernatant.
Xylan reaction products were identified as xylobiose, xylotriose, and xylotetraose. K
m values (mg ml−1) for soluble oat spelt and birchwood xylan were 11.8 and 9.45, respectively. Although PXII-1 showed 85% activity retention
upon incubation at 50°C and pH 5.0 for 20 days, incubation at pH 7.0 resulted in 50% activity loss within 3 days. PXII-1 stability
at pH 7.0 was improved in the presence of 20 mM cysteine, which allowed for 85% activity retention for 25 days. This study
on the production in high yields of a remarkably thermostable xylanase is of significance due to the central role that this
class of biocatalyst shares, along with cellulases, for the much needed enzymatic hydrolysis of biomass. Furthermore, stable
xylanases are important for the manufacture of paper, animal feed, and xylooligosaccharides. 相似文献
20.
Amal Hammami Ahmed Bayoudh Bilel Hadrich Ola Abdelhedi Mourad Jridi Moncef Nasri 《Biotechnology progress》2020,36(3):e2965
This work deals with the optimization of the culture conditions of Bacillus invictae AH1 in order to increase the production level of the proteolytic activity. Response-surface methodology (RSM) was applied for the most significant fermentation parameters (concentration of wheat bran and K2HPO4/KH2PO4) that were earlier identified by Plackett–Burman Design from seven possible factors. A central composite design was used and the quadratic regression model of producing active protease was built. A maximum protease activity was reached and validated experimentally, using a maximum wheat bran concentration (50 g/L) with increased K2HPO4/KH2PO4 concentration (2.275 g/L). Protease production obtained experimentally coincident with the predicted value and the model was proven to be adequate. Interestingly, the use of RSM increased the protease production by four times (7,000 U/mL) using a low-cost substrate and a culture time of 40 hr, as compared to the standard culture conditions. In the second part of this study, a H2O2-tolerant alkaline protease produced from B. invictae AH1 with a molecular mass of about 41 kDa, noted P3, was purified by successive steps of ultrafiltration, gel filtration and ion exchange chromatography. The K m and Vmax values of the purified protease using casein, as substrate, were about 4 mg/mL and 27 μM/min, respectively. The highest enzyme activity was found at pH 9.0 and a temperature of 60°C. In addition, the enzyme showed a quasi-total stability against H2O2 (5% for 1 hr) and against most of the tested solid and liquid detergents, suggesting its eventual use in bio-detergent formulations. 相似文献