scholarly journals Production of Ethanol from Hemicellulosic Sugars of Exhausted Olive Pomace by Escherichia coli

Processes ◽  
2020 ◽  
Vol 8 (5) ◽  
pp. 533 ◽  
Author(s):  
Juan Carlos López-Linares ◽  
Irene Gómez-Cruz ◽  
Encarnación Ruiz ◽  
Inmaculada Romero ◽  
Eulogio Castro
Keyword(s):  
Escherichia Coli ◽  
Activated Charcoal ◽  
Sugar Solution ◽  
Olive Pomace ◽  
Acid Pretreatment ◽  
Fermentation Time ◽  
E Coli ◽  
Inhibitory Compounds ◽  
Ethanol Yields ◽  
Hemicellulosic Sugars

Exhausted olive pomace (EOP) is the main residue generated in olive oil industries, after the extraction of the residual oil from olive pomace with hexane. This work studies the ethanol production from hemicellulosic sugars of EOP. The fermentability of the sugar solution, resulting from the acid pretreatment of EOP, was evaluated using Escherichia coli SL100, although a detoxification step was required before fermentation. Overliming and activated charcoal detoxification were tested to minimize the presence of inhibitory compounds in the hydrolysate and to achieve a fermentable medium. E. coli assimilated all sugars in both detoxified hydrolysates and achieved ethanol yields of about 90% of the theoretical one. However, the fermentation time was much shorter when the hydrolysate had been detoxified with activated charcoal (20 h versus 120 h).

scholarly journals OPTIMASI PROSES UNTUK EKSPRESI GEN ENDOGLUKANASE DARI Bacillus sp. RP1 OLEH Escherichia coli BL21 (DE3)/ egc

2018 ◽  
Vol 5 (1) ◽  
pp. 44
Author(s):  
Hans Victor ◽  
Maelita Ramdani Moeis
Keyword(s):  
Escherichia Coli ◽  
Enzyme Activity ◽  
Process Optimization ◽  
Rice Hull ◽  
Bacillus Sp ◽  
Fermentation Time ◽  
E Coli ◽  
Cell Weight ◽  
Dry Cell Weight ◽  
Dry Cell

Process Optimization for Endoglucanase Gene Expression Derived from Bacillus sp. RP1 by Escherichia coli BL21 (DE3)/egcABSTRACTCellulases are one of the most used enzymes in industrial processes. In an effort to increase production, industries have developed strategies such as isolating new cellulase producing strains, genetic engineering and process optimization since the last 50 years. One endoglucanase producing strain, Bacillus sp. RP1 was isolated from hot springs. The ribosome binding site and coding sequence of the endoglucanase gene (egc) from Bacillus sp. RP1 was cloned into pGEM-T Easy. The recombinant plasmid was used to transform E. coli BL21 (DE3). Cloning was followed by process optimization. Medium composition was selected using Plackett-Burman design. The medium components tested were rice hull, molasses, ammonium chloride, urea and fishmeal. Rice hull and molasses were found to be the factors most influencing enzyme activity and dry cell weight, respectively. The next step involved Box-Behnken method and response surface methodology to optimize the responses against molasses concentration, rice hull concentration and fermentation time. The concentration intervals used to test were 1%, 5.5% and 10% while the fermentation time used were 24, 36 and 48 hours. The conditions which optimized both enzyme activity and dry cell weight were 7.45% molasses, 6.45% rice hull and 39.52 hours of fermentation.Keywords: Bacillus sp. RP1, E. coli BL21 (DE3), egc, Endoglucanase, optimization ABSTRAKSelulase adalah salah satu enzim yang banyak dimanfaatkan dalam berbagai industri. Sebagai upaya untuk memenuhi kebutuhan, 50 tahun terakhir dikembangkan beberapa strategi untuk meningkatkan produksi selulase yang mencakup rekayasa genetika dan optimasi proses. Karena itu, dilakukan kloning gen egc dan RBS yang berasal dari Bacillus sp. RP1 yang diisolasi dari sumber air panas ke dalam vektor pGEM-T Easy. E. coli BL21 (DE3) ditransformasikan dengan vektor yang mengandung gen egc tersebut. Setelah kloning, optimasi proses berupa desain medium turut dilakukan untuk mengoptimalkan ekspresi gen egc. Desain medium diawali dengan seleksi komposisi medium menggunakan metode Plackett-Burman. Komponen medium yang diuji adalah kulit beras, molase, amonium klorida, urea dan tepung ikan. Kulit beras dan molase diperoleh sebagai bahan yang paling berpengaruh terhadap aktivitas enzim dan berat kering sel. Tahap selanjutnya melibatkan metode statistik Box-Behnken dan metodologi respons permukaan yang bertujuan mengoptimalkan respons aktivitas enzim dan berat kering sel terhadap konsentrasi molase, konsentrasi kulit beras dan lama fermentasi. Konsentrasi yang diuji adalah 1%, 5,5% dan 10%, sedangkan lama fermentasi yang diuji adalah 24, 36 dan 48 jam. Konsentrasi optimal molase adalah 7,45% dan konsentrasi optimal kulit beras adalah 6,45% dengan lama fermentasi optimal 39,52 jam.Kata Kunci: Bacillus sp. RP1, E. coli BL21 (DE3), egc, Endoglukanase, optimasi

scholarly journals Highly Efficient Biotransformation of Eugenol to Ferulic Acid and Further Conversion to Vanillin in Recombinant Strains of Escherichia coli

2003 ◽  
Vol 69 (11) ◽  
pp. 6569-6576 ◽  
Author(s):  
Jörg Overhage ◽  
Alexander Steinbüchel ◽  
Horst Priefert
Keyword(s):  
Escherichia Coli ◽  
Ferulic Acid ◽  
Alcohol Oxidase ◽  
Coniferyl Alcohol ◽  
Vanillyl Alcohol ◽  
Resting Cells ◽  
Fermentation Time ◽  
Molar Yield ◽  
E Coli ◽  
Maximum Production Rate

ABSTRACT The vaoA gene from Penicillium simplicissimum CBS 170.90, encoding vanillyl alcohol oxidase, which also catalyzes the conversion of eugenol to coniferyl alcohol, was expressed in Escherichia coli XL1-Blue under the control of the lac promoter, together with the genes calA and calB, encoding coniferyl alcohol dehydrogenase and coniferyl aldehyde dehydrogenase of Pseudomonas sp. strain HR199, respectively. Resting cells of the corresponding recombinant strain E. coli XL1-Blue(pSKvaomPcalAmcalB) converted eugenol to ferulic acid with a molar yield of 91% within 15 h on a 50-ml scale, reaching a ferulic acid concentration of 8.6 g liter−1. This biotransformation was scaled up to a 30-liter fermentation volume. The maximum production rate for ferulic acid at that scale was 14.4 mmol per h per liter of culture. The maximum concentration of ferulic acid obtained was 14.7 g liter−1 after a total fermentation time of 30 h, which corresponded to a molar yield of 93.3% with respect to the added amount of eugenol. In a two-step biotransformation, E. coli XL1-Blue(pSKvaomPcalAmcalB) was used to produce ferulic acid from eugenol and, subsequently, E. coli(pSKechE/Hfcs) was used to convert ferulic acid to vanillin (J. Overhage, H. Priefert, and A. Steinbüchel, Appl. Environ. Microbiol. 65:4837-4847, 1999). This process led to 0.3 g of vanillin liter−1, besides 0.1 g of vanillyl alcohol and 4.6 g of ferulic acid liter−1. The genes ehyAB, encoding eugenol hydroxylase of Pseudomonas sp. strain HR199, and azu, encoding the potential physiological electron acceptor of this enzyme, were shown to be unsuitable for establishing eugenol bioconversion in E. coli XL1-Blue.

High-Throughput Small Molecule Screening Reveals Structurally Diverse Compounds That Inhibit the Growth of Escherichia coli O157:H7 In Vitro

2011 ◽  
Vol 74 (12) ◽  
pp. 2148-2156
Author(s):  
J. C. CHEN ◽  
B. A. CARLSON ◽  
J. N. SOFOS ◽  
G. C. SMITH ◽  
K. E. BELK ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Hemolytic Uremic Syndrome ◽  
Small Molecule ◽  
High Throughput ◽  
Food Supply ◽  
Escherichia Coli O157 ◽  
Human Food ◽  
E Coli ◽  
Inhibitory Compounds ◽  
Uremic Syndrome

Escherichia coli O157:H7 colonizes the gastrointestinal tract of ruminants asymptomatically and may enter the human food supply through fecal contamination. A fraction of individuals infected by E. coli O157:H7 develop hemolytic uremic syndrome, a life-threatening condition. When individuals infected by E. coli O157:H7 are treated with certain antibiotics, an increased incidence of hemolytic uremic syndrome may result. This finding supports the need to identify novel compounds that can either reduce the load of E. coli O157:H7 entering the human food supply or serve as alternative therapeutic treatments for infected individuals. We developed a high-throughput turbidometric assay to identify novel compounds that inhibit E. coli O157:H7 growth. Pin transfers were performed to introduce small molecule libraries into 384-well plates, where each well contained approximately 5.0 log CFU of E. coli O157:H7. Plates were incubated at 37°C for 18 h, and the optical density was measured to determine the effect of each small molecule. A total of 64,562 compounds were screened in duplicate, and 43 unique compounds inhibited E. coli O157:H7 growth. Thirty-eight of the 43 inhibitory compounds belonged to known bioactive libraries, and the other 5 compounds were from commercial libraries derived from splitting and pooling. Inhibitory compounds from known bioactive libraries were most frequently therapeutic antibiotics (n = 34) but also included an antiviral compound, a compound that disrupts the citric acid cycle, and two biguanide compounds, which have been used for various nonclinical applications. We identified two novel compounds (i.e., biguanides) that should be studied further for their ability to reduce pathogen populations in foods.

scholarly journals Microbial Conversion of Glycerol to 1,3-Propanediol by an Engineered Strain of Escherichia coli

2009 ◽  
Vol 75 (6) ◽  
pp. 1628-1634 ◽  
Author(s):  
Xueming Tang ◽  
Yongsong Tan ◽  
Hong Zhu ◽  
Kai Zhao ◽  
Wei Shen
Keyword(s):  
Escherichia Coli ◽  
Temperature Shift ◽  
Sole Source ◽  
Temperature Sensitive ◽  
Lambda Phage ◽  
Microbial Conversion ◽  
Two Stage ◽  
Fermentation Time ◽  
E Coli ◽  
K 12

ABSTRACT In an effort to improve industrial production of 1,3-propanediol (1,3-PD), we engineered a novel polycistronic operon under the control of the temperature-sensitive lambda phage PLPR promoter regulated by the cIts857 repressor and expressed it in Escherichia coli K-12 ER2925. The genes for the production of 1,3-PD in Clostridium butyricum, dhaB1 and dhaB2, which encode the vitamin B12-independent glycerol dehydratase DhaB1 and its activating factor, DhaB2, respectively, were tandemly arrayed with the E. coli yqhD gene, which encodes the 1,3-propanediol oxidoreductase isoenzyme YqhD, an NADP-dependent dehydrogenase that can directly convert glycerol to 1,3-PD. The microbial conversion of 1,3-PD from glycerol by this recombinant E. coli strain was studied in a two-stage fermentation process. During the first stage, a novel high-cell-density fermentation step, there was significant cell growth and the majority of the metabolites produced were organic acids, mainly acetate. During the second stage, glycerol from the fresh medium was rapidly converted to 1,3-PD following a temperature shift from 30°C to 42°C. The by-products were mainly pyruvate and acetate. During this two-stage process, the overall 1,3-PD yield and productivity reached 104.4 g/liter and 2.61 g/liter/h, respectively, and the conversion rate of glycerol to 1,3-PD reached 90.2% (g/g). To our knowledge, this is the highest reported yield and productivity efficiency of 1,3-PD with glycerol as the sole source of carbon. Furthermore, the overall fermentation time was only 40 h, shorter than that of any other reports.

Sites of Adsorption of the Bacteriophages T3 and T5 on the Wall of Escherichia Coli B.

1967 ◽  
Vol 25 ◽  
pp. 96-97
Author(s):  
Manfred E. Bayer
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Electron Microscope ◽  
Uranyl Acetate ◽  
E Coli ◽  
Receptor Sites ◽  
Rigid Cell Wall ◽  
Host Cell Surface ◽  
Bacterial Viruses ◽  
Escherichia Coli B

Bacterial viruses adsorb specifically to receptors on the host cell surface. Although the chemical composition of some of the cell wall receptors for bacteriophages of the T-series has been described and the number of receptor sites has been estimated to be 150 to 300 per E. coli cell, the localization of the sites on the bacterial wall has been unknown.When logarithmically growing cells of E. coli are transferred into a medium containing 20% sucrose, the cells plasmolize: the protoplast shrinks and becomes separated from the somewhat rigid cell wall. When these cells are fixed in 8% Formaldehyde, post-fixed in OsO4/uranyl acetate, embedded in Vestopal W, then cut in an ultramicrotome and observed with the electron microscope, the separation of protoplast and wall becomes clearly visible, (Fig. 1, 2). At a number of locations however, the protoplasmic membrane adheres to the wall even under the considerable pull of the shrinking protoplast. Thus numerous connecting bridges are maintained between protoplast and cell wall. Estimations of the total number of such wall/membrane associations yield a number of about 300 per cell.

The Influence of Glycosylation on the Catalytic and Fibrinolytic Properties of Pro-Urokinase

1992 ◽  
Vol 68 (05) ◽  
pp. 539-544 ◽  
Author(s):  
Catherine Lenich ◽  
Ralph Pannell ◽  
Jack Henkin ◽  
Victor Gurewich
Keyword(s):  
Escherichia Coli ◽  
Mammalian Cell ◽  
Human Gene ◽  
Culture Media ◽  
Mammalian Cell Culture ◽  
Glycosylation Site ◽  
Clot Lysis ◽  
Cell Culture Media ◽  
E Coli ◽  
The Uk

SummaryWe previously found that human pro-UK expressed in Escherichia coli is more active in fibrinolysis than recombinant human pro-UK obtained from mammalian cell culture media. To determine whether this difference is related to the lack of glycosylation of the E. coli product, we compared the activity of E. coli-derived pro-UK [(-)pro-UK] with that of a glycosylated pro-UK [(+)pro-UK] and of a mutant of pro-UK missing the glycosylation site at Asn-302 [(-) (302) pro-UK]. The latter two pro-UKs were obtained by expression of the human gene in a mammalian cell. The nonglycosylated pro-UKs were activated by plasmin more efficiently (≈2-fold) and were more active in clot lysis (1.5-fold) than the (+)pro-UK. Similarly, the nonglycosylated two-chain derivatives (UKs) were more active against plasminogen and were more rapidly inactivated by plasma inhibitors than the (+)UK.These findings indicate that glycosylation at Asn-302 influences the activity of pro-UK/UK and could be the major factor responsible for the enhanced activity of E. coli-derived pro-UK.

Usefulness of a Multiplex PCR for Detection of Diarrheagenic Escherichia coli in a Diagnostic Microbiology Laboratory Setting

2016 ◽  
Vol 1 (2) ◽  
pp. 38-42 ◽  
Author(s):  
Khairun Nessa ◽  
Dilruba Ahmed ◽  
Johirul Islam ◽  
FM Lutful Kabir ◽  
M Anowar Hossain
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Clinical Laboratory ◽  
Proteinase K ◽  
Microbiology Laboratory ◽  
Pcr Assay ◽  
E Coli ◽  
Diarrheagenic Escherichia Coli ◽  
Multiplex Pcr Assay ◽  
Diagnostic Microbiology

A multiplex PCR assay was evaluated for diagnosis of diarrheagenic Escherichia coli in stool samples of patients with diarrhoea submitted to a diagnostic microbiology laboratory. Two procedures of DNA template preparationproteinase K buffer method and the boiling method were evaluated to examine isolates of E. coli from 150 selected diarrhoeal cases. By proteinase K buffer method, 119 strains (79.3%) of E. coli were characterized to various categories by their genes that included 55.5% enteroaggregative E. coli (EAEC), 18.5% enterotoxigenic E. coli (ETEC), 1.7% enteropathogenic E. coli (EPEC), and 0.8% Shiga toxin-producing E. coli (STEC). Although boiling method was less time consuming (<24 hrs) and less costly (<8.0 US $/ per test) but was less efficient in typing E. coli compared to proteinase K method (41.3% vs. 79.3% ; p<0.001). The sensitivity and specificity of boiling method compared to proteinase K method was 48.7% and 87.1% while the positive and negative predictive value was 93.5% and 30.7%, respectively. The majority of pathogenic E. coli were detected in children (78.0%) under five years age with 53.3% under one year, and 68.7% of the children were male. Children under 5 years age were frequently infected with EAEC (71.6%) compared to ETEC (24.3%), EPEC (2.7%) and STEC (1.4%). The multiplex PCR assay could be effectively used as a rapid diagnostic tool for characterization of diarrheagenic E. coli using a single reaction tube in the clinical laboratory setting.Bangladesh J Med Microbiol 2007; 01 (02): 38-42

Antagonistic activity of Bacillus subtilis strains isolated from various sources

2018 ◽  
Vol 8 (2) ◽  
pp. 354-364
Author(s):  
A. N. Irkitova ◽  
A. V. Grebenshchikova ◽  
A. V. Matsyura
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Wild Strain ◽  
Fish Farming ◽  
Antagonistic Activity ◽  
Antagonistic Effect ◽  
E Coli ◽  
Long Period ◽  
Test Culture ◽  
Agar Media

<p>An important link in solving the problem of healthy food is the intensification of the livestock, poultry and fish farming, which is possible only in the adoption and rigorous implementation of the concept of rational feeding of animals. In the implementation of this concept required is the application of probiotic preparations. Currently, there is an increased interest in spore probiotics. In many ways, this can be explained by the fact that they use no vegetative forms of the bacilli and their spores. This property provides spore probiotics a number of advantages: they are not whimsical, easily could be selected, cultivated, and dried. Moreover, they are resistant to various factors and could remain viable during a long period. One of the most famous spore microorganisms, which are widely used in agriculture, is <em>Bacillus subtilis</em>. Among the requirements imposed to probiotic microorganisms is mandatory – antagonistic activity to pathogenic and conditional-pathogenic microflora. The article presents the results of the analysis of antagonistic activity of collection strains of <em>B. subtilis</em>, and strains isolated from commercial preparations. We studied the antagonistic activity on agar and liquid nutrient medias to trigger different antagonism mechanisms of <em>B. subtilis</em>. On agar media, we applied three diffusion methods: perpendicular bands, agar blocks, agar wells. We also applied the method of co-incubating the test culture (<em>Escherichia coli</em>) and the antagonist (or its supernatant) in the nutrient broth. Our results demonstrated that all our explored strains of <em>B. subtilis</em> have antimicrobial activity against a wild strain of <em>E. coli</em>, but to varying degrees. We identified strains of <em>B. subtilis</em> with the highest antagonistic effect that can be recommended for inclusion in microbial preparations for agriculture.</p><p><em><br /></em><em></em></p>

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