| 11. |
- Birgisson, Hakon, et al.
(författare)
-
Two new thermostable alpha-L-rhamnosidases from a novel thermophilic bacterium
- 2004
-
Ingår i: Enzyme and Microbial Technology. - : Elsevier BV. - 0141-0229. ; 34:6, s. 561-571
-
Tidskriftsartikel (refereegranskat)abstract
- Two new thermostable alpha-L-rhamnosidases with novel substrate hydrolysis pattern were cloned and expressed from a new thermophilic bacterium. Fragments of the two alpha-L-rhamnosidase genes, rhmA and rhmB were identified in a partially sequenced genome of the bacterium. Whole genes were recovered by amplifying flanking sequences with single specific primers and nonspecific walking primers. The recovered Genes were then cloned into Escherichia coli and their enzymes produced and purified. Both enzymes were dimers and the MW of the monomers. were 104 and 107 kDa for RhmA and RhmB, respectively. Both rhamnosidases had a temperature optimum at 70degreesC. RhmA had pH optimum at 7.9 and RhmB had a broad pH optimum of 5.0 to 6.9 and RhmA had over 50% activity in the pH interval 5.0 to 8.7 and RhmB in the pH interval 4.0 to 7.9. Both enzymes had over 20% residual activity after 24-h incubation at 60degreesC. RhmA and RhmB had K values of 0.46 and 0.66 mM and V-max values of 134 and 352 U mg(-1) respectively, on p-nitrophenyl-alpha-L-rhamnopyrano side. Both rhamnosidases were active on both alpha-1,2- and alpha-1,6-linkages to beta-D-glucoside. (C) 2004 Elsevier Inc. All rights reserved.
|
|
| 12. |
- Björnsson, Lovisa, et al.
(författare)
-
Evaluation of new methods for the monitoring of alkalinity dissolved hydrogen and the microbial community in anaerobic digestion
- 2001
-
Ingår i: Water Research. - 1879-2448. ; 35:12, s. 2833-2840
-
Tidskriftsartikel (refereegranskat)abstract
- New methods for spectrophotometric alkalinity measurement, dissolved hydrogen monitoring and for obtaining a fingerprint of the microbial community were evaluated as tools for process monitoring in anaerobic digestion. The anaerobic digestion process was operated at organic loading rates of 1.5, 3.0 and 4.5g volatile solids l-1d-1 and subjected to pulse loads of carbohydrate, lipid, protein and a mixed sludge substrate. The spectrophotometric alkalinity monitoring method showed good agreement with traditional titrimetric alkalinity monitoring and has the advantage of being easy to modify to on-line monitoring applications. The on-line monitoring of dissolved hydrogen gave valuable information about approaching process overload and can be a good complement to the conventional monitoring of volatile fatty acids. Changing process conditions were also reflected in the microbial fingerprint that could be achieved by partitioning in two-phase systems. The investigated methods showed potential for application in increasing our understanding of the anaerobic digestion process as well as for being applicable for monitoring in the complex environment of full-scale anaerobic digestion processes.
|
|
| 13. |
- Björnsson, Lovisa, et al.
(författare)
-
Evaluation of parameters for monitoring an anaerobic co-digestion process
- 2000
-
Ingår i: Applied Microbiology and Biotechnology. - : Springer Science and Business Media LLC. - 1432-0614 .- 0175-7598. ; 54:6, s. 844-849
-
Tidskriftsartikel (refereegranskat)abstract
- The system investigated in this study is an anaerobic digester at a municipal wastewater treatment plant operating on sludge from the wastewater treatment, co-digested with carbohydrate-rich food-processing waste. The digester is run below maximum capacity to prevent overload. Process monitoring at present is not extensive, even for the measurement of on-line gas production rate and off-line pH. Much could be gained if a better program for monitoring and control was developed, so that the full capacity of the system could be utilised without the risk of overload. The only limit presently set for correct process operation is that the pH should be above 6.8. In the present investigation, the pH was compared with alkalinity, gas production rate, gas composition and the concentration of volatile fatty acids (VFA). Changes in organic load were monitored in the full-scale anaerobic digester and in laboratory-scale models of the plant. Gas-phase parameters showed a slow response to changes in load. The VFA concentrations were superior for indicating overload of the microbial system, but alkalinity and pH also proved to be good monitoring parameters. The possibility of using pH as a process indicator is, however, strongly dependent on the buffering capacity. In this study, a minor change in the amount of carbohydrates in the substrate had drastic effects on the buffering effect of the system.
|
|
| 14. |
- Björnsson, Lovisa, et al.
(författare)
-
Utilisation of a Pd-MOS sensor for on-line monitoring of dissolved hydrogen in anaerobic digestion
- 2001
-
Ingår i: Biotechnology and Bioengineering. - 1097-0290. ; 73:1, s. 35-43
-
Tidskriftsartikel (refereegranskat)abstract
- The use of a hydrogen-sensitive palladium-metal oxide semiconductor (Pd-MOS) sensor in combination with a membrane for liquid-to-gas transfer for the detection of dissolved hydrogen was investigated. The system was evaluated with known concentrations of dissolved hydrogen in water. The lowest concentration detected with this set-up was 160 nM. The method was applied to monitoring of a laboratory-scale anaerobic digestion process employing mixed sludge containing mainly food/industrial waste. Pulse loads of glucose were added to the system at different levels of microbial activity, and the microbial status of the culture was reflected in the dissolved hydrogen response. Simultaneous headspace hydrogen measurements were performed, and at the lower levels of dissolved hydrogen no corresponding headspace hydrogen could be detected. When glucose was added to a resting culture the dissolved hydrogen response was rapid and the first response could be detected 9 min after addition of glucose, whereas headspace hydrogen concentrations increased only after 80 to 110 min. This indicates limitations in the liquid-to-gas hydrogen transfer and illustrates the importance of hydrogen monitoring in the liquid. The sensor system developed is flexible, the membrane is easily replaceable, and the probe for liquid-to-gas hydrogen transfer can be adjusted easily to large-scale applications.
|
|
| 15. |
- Borde, X, et al.
(författare)
-
Synergistic relationships in algal-bacterial microcosms for the treatment of aromatic pollutants.
- 2003
-
Ingår i: Bioresource Technology. - 1873-2976. ; 86:3, s. 293-300
-
Tidskriftsartikel (refereegranskat)abstract
- The potential of algal–bacterial microcosms was studied for the biodegradation of salicylate, phenol and phenanthrene. The isolation and characterization of aerobic bacterial strains capable of mineralizing each pollutant were first conducted. Ralstonia basilensis was isolated for salicylate degradation, Acinetobacter haemolyticus for phenol and Pseudomonas migulae and Sphingomonas yanoikuyae for phenanthrene. The green alga Chlorella sorokiniana was then cultivated in the presence of the pollutants at different concentrations, showing increasing inhibitory effects in the following order: salicylate85%) was recorded only in the systems inoculated with both algae and bacteria and incubated under continuous lighting. This study presents, to our knowledge, the first reported case of photosynthesis-enhanced biodegradation of toxic aromatic pollutants by algal–bacterial microcosms in a one-stage treatment.
|
|
| 16. |
- Bradoo, Sapna, et al.
(författare)
-
Synthesis of alkylgalactosides using whole cells of Bacillus pseudofirmus species as catalysts
- 2004
-
Ingår i: Journal of Biotechnology. - : Elsevier BV. - 1873-4863 .- 0168-1656. ; 110:3, s. 273-285
-
Tidskriftsartikel (refereegranskat)abstract
- Whole cells of alkaliphilic Bacillus pseudofirmus AR-199, induced for beta-galactosidase activity, were used for the synthesis of 1-hexyl-beta-D-galactoside and 1-octyl-beta-D-galactoside, respectively, by transglycosylation reaction between lactose and the corresponding alcohol acceptor. The product yield was strongly influenced by the initial water content in the reaction mixture. Water content of 10% (v/v) was optimal providing 3.6-36 mM hexyl galactoside from 10 to 150 mM lactose, and no secondary product hydrolysis. Product yield could be enhanced by supplementing the reaction mixture with more cells or partly replacing the product with fresh substrate, but was decreased with time to the initial equilibrium level. Cell permeabilisation or disruption resulted in increased reaction rate and higher product yield but was followed by product hydrolysis. Octyl galactoside synthesis using whole cells was optimal at water content of 2% (v/v) with a yield of 26%. The cells were immobilised in cryogels of polyvinyl alcohol for use in continuous process, where hexyl galactoside was produced with a constant yield of 50% from 50 mM lactose for at least a week. (C) 2004 Elsevier B.V. All rights reserved.
|
|
| 17. |
- Dainiak, Maria, et al.
(författare)
-
Direct capture of product from fermentation broth using a cell-repelling ion exchanger.
- 2002
-
Ingår i: Journal of Chromatography A. - 0021-9673. ; 942:1-2, s. 123-131
-
Tidskriftsartikel (refereegranskat)abstract
- A new technique for treating anion exchangers has been proposed allowing direct capture of the fermentation product, shikimic acid directly from the cell-containing fermentation broth. A layer of hydrophilic polymer, poly(acrylic acid) (PAA) has been physically adsorbed on the anion exchanger followed by a covalent cross-linking of PAA. The PAA layer is penetrable for small molecules despite being negatively charged as PAA is, but the polymer layer repels large negatively charged structures like cell debris and cells preventing them from adsorption to the chromatographic matrix. The binding capacity for pure shikimic was about 81 mg/ml adsorbent for both cross-linked PAA-Amberlite and native Amberlite in the fluidized mode of column operation. Binding capacity dropped to 17 and 15 mg per ml adsorbent, respectively, when using filtrated fermentation broth and to about 10 mg/ml adsorbent for cross-linked PAA-Amberlite when using directly the fermentation broth containing cells. Native Amberlite cannot be used for the direct capture of shikimic acid due to the immediate clogging of the column and the collapse of the expanded bed. The cross-linked PAA-Amberlite was used repeatedly for the direct adsorption of shikimic acid from the industrial fermentation broth.
|
|
| 18. |
- Dainiak, Maria, et al.
(författare)
-
Integrated isolation of antibody fragments from microbial cell culture fluids using supermacroporous cryogels
- 2004
-
Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1045:1-2, s. 93-98
-
Tidskriftsartikel (refereegranskat)abstract
- The present paper describes a chromatographic capture/purification step for the recovery of proteins directly from undiluted and unclarified cell culture broths using supermacroporous dimethylacrylamide (DMAA) cryogel. The interconnected character and the size (10-100 mum) of the pores of the adsorbent make it possible to process whole cell fermentation broths without blocking the column. Cu2+-iminodiacetic acid (IDA) DMAA cryogel has been used for the isolation and purification of excreted (His)(6)-tagged single chain (sc) Fv antibody fragments, (His)(6)-scFv, from E. coli cell culture. Bound protein was recovered with 0.2 M imidazole or with 20 mM EDTA and was practically cell-free. Chromatographic capture using Cu2+-IDA cryogel column was performed at flow rates of 300 and 600 cm/h, respectively and resulted in 84-96% recovery of (His)(6)-scFv fragments with a purification factor of 13-15. The DMAA cryogel adsorbent is mechanically stable, can withstand harsh cleaning-in-place procedure and is relatively inexpensive. Chromatographic isolation of proteins using cryogels allows efficient removal of cells and can be operated at a flow rate as high as 600 cm/h. This novel technique has proven to be a scalable process, does not require special equipment and can be a good alternative to expanded bed adsorption and other integrated isolation techniques. (C) 2004 Published by Elsevier B.V.
|
|
| 19. |
- Dainiak, Maria, et al.
(författare)
-
Polyelectrolyte-coated ion exchangers for cell-resistant expanded bed adsorption
- 2002
-
Ingår i: Biotechnology Progress. - : Wiley. - 1520-6033 .- 8756-7938. ; 18:4, s. 815-820
-
Tidskriftsartikel (refereegranskat)abstract
- Adsorption chromatography in expanded beds is a widely used technology for direct capture of target proteins from fermentation broths. However, in many cases this method cannot be applied as a result of the strong tendency of cells or cell debris to interact with the adsorbent beads. To prevent contamination of the expanded bed with the biomass, STREAMLINE DEAE, anion exchanger designed for expanded bed adsorption, was modified with a layer of poly(acrylic acid) (PAA). The shielding layer of polyelectrolyte was attached to the surface of the matrix beads via electrostatic interactions. PAA with a high degree of polymerization was chosen to prevent diffusion of large polymer molecules into the pores of adsorbent. Thus, the shielding layer of PAA was adsorbed, only at the mouth of the pores of STREAMLINE DEAE beads and only marginally decreased the binding capacity of the ion exchanger for bovine serum albumin, the model protein in this study. PAA-coated STREAMLINE DEAE practically did not interact with yeast cells, which other-wise bound,strongly to the native adsorbent at neutral conditions. Cell-resistant PAA-coated anion exchanger was successfully used for isolation of BSA from the model protein mixture containing BSA, lysozyme (positively charged at applied conditions), and yeast cells. The layer of PAA was stable under mild elution conditions, and the modified adsorbent could be used in the repeated purification cycles.
|
|
| 20. |
- Fernandes, Sheryl, et al.
(författare)
-
Recovery of recombinant cutinase using detergent foam
- 2002
-
Ingår i: Biotechnology Progress. - : Wiley. - 1520-6033 .- 8756-7938. ; 18:1, s. 116-123
-
Tidskriftsartikel (refereegranskat)abstract
- Foam generated by vigorous stirring of a nonionic detergent, Triton X-114, was used for the recovery of recombinant cutinase expressed by Saccharomyces cerevisiae. The enzyme with a hydrophobic fusion tag, (Trp-Pro)(4), was recovered with a higher yield as compared to the wild-type cutinase, indicating the involvement of hydrophobic interactions in protein isolation with the foam. The influence of various factors including volume, dilution, pH, different additives, and cell concentration in the medium on enzyme recovery was investigated. Interaction of the enzyme with detergent was monitored using fluorescence spectroscopy. No significant changes in protein conformation after the isolation procedure were observed using circular dichroism.
|
|
