| 1. |
- Ademark, Pia, et al.
(author)
-
Hydrolytic properties of a beta-mannosidase purified from Aspergillus niger. J. Biotechnol. 75: 281-289.
- 1999
-
In: Journal of Biotechnology. - 1873-4863. ; 75:2-3, s. 281-289
-
Journal article (peer-reviewed)abstract
- A β-mannosidase was purified to homogeneity from the culture filtrate of Aspergillus niger. A specific activity of 500 nkat mg−1 and a 53-fold purification was achieved using ammonium sulfate precipitation, anion-exchange chromatography, and gel filtration. The isolated enzyme has an isoelectric point of 5.0 and appears to be a dimer composed of two 135-kDa subunits. It is a glycoprotein and contains 17% N-linked carbohydrate by weight. Maximal activity was observed at pH 2.4–5.0 and at 70°C. The β-mannosidase hydrolyzed β-1,4-linked manno-oligosaccharides of degree of polymerization (DP) 2–6 and also released mannose from polymeric ivory nut mannan and galactomannan. The Km and Vmax values for p-nitrophenyl-β-Image-mannopyranoside were 0.30 mM and 500 nkat mg−1, respectively. Hydrolysis of Image-galactose substituted manno-oligosaccharides showed that the β-mannosidase was able to cleave up to, but not beyond, a side group. An internal peptide sequence of 15 amino acids was highly similar to that of an Aspergillus aculeatus β-mannosidase belonging to family 2 of glycosyl hydrolases.
|
|
| 2. |
- Ademark, Pia, et al.
(author)
-
Softwood hemicellulose-degrading enzymes from Aspergillus niger: Purification and properties of a β-mannanase
- 1998
-
In: Journal of Biotechnology. - 1873-4863. ; 63:3, s. 199-210
-
Journal article (peer-reviewed)abstract
- The enzymes needed for galactomannan hydrolysis, i.e. β-mannanase, α-galactosidase and β-mannosidase, were produced by the filamentous fungus Aspergillus niger. The β-mannanase was purified to electrophoretic homogeneity in three steps using ammonium sulfate precipitation, anion-exchange chromatography and gel filtration. The purified enzyme had an isoelectric point of 3.7 and a molecular mass of 40 kDa. Ivory nut mannan was degraded mainly to mannobiose and mannotriose when incubated with the β-mannanase. Analysis by 1H NMR spectroscopy during hydrolysis of mannopentaose showed that the enzyme acts by the retaining mechanism. The N-terminus of the purified A. niger β-mannanase was sequenced by Edman degradation, and comparison with Aspergillus aculeatus β-mannanase indicated high identity. The enzyme most probably lacks a cellulose binding domain since it was unable to adsorb on cellulose.
|
|
| 3. |
- Ahlqvist, Josefin, et al.
(author)
-
Affinity binding of inclusion bodies on supermacroporous monolithic cryogels using labeling with specific antibodies
- 2006
-
In: Journal of Biotechnology. - : Elsevier BV. - 0168-1656 .- 1873-4863. ; 122:2, s. 216-225
-
Journal article (peer-reviewed)abstract
- A new chromatographic method based on affinity supermacroporous monolithic cryogels is developed for binding and analyzing inclusion bodies during fermentation. The work demonstrated that it is possible to bind specific IgG and IgY antibodies to the 15 and 17 amino acids at the terminus ends of a 33 kDa target protein aggregated as inclusion bodies. The antibody treated inclusion bodies from lysed fermentation broth can be specifically retained in protein A and pseudo-biospecific ligand sulfamethazine modified supermacroporous cryogels. The degree of binding of IgG and IgY treated inclusion bodies to the Protein A and sulfamethazine gels are investigated, as well as the influence of pH on the sulfamethazine ligand. Optimum binding of 78 and 72% was observed on both protein A and sulfamethazine modified cryogel columns, respectively, using IgG labeling of the inclusion bodies. The antibody treated inclusion bodies pass through unretained in the sulfamethazine supermacroporous gel at pH that does not favour the binding between the ligand on the gel and the antibodies on the surface of inclusion bodies. Also the unlabeled inclusion bodies went through the gel unretained, showing no non-specific binding or trapping within the gel. These findings may very well be the foundation for the building of a powerful analytical tool during fermentation of inclusion bodies as well as a convenient way to purify them from fermentation broth. These results also support our earlier findings [Kumar, A., Plieva, F.M., Galaev, I.Yu., Mattiasson, B.. 2003. Affinity fractionation of lymphocytes using a monolithic cyogel. J. Immunol. Methods 283, 185-194] with mammalian cells that were surface labeled with specific antibodies and recognized on protein A supermacroporous gels. A general binding and separation system can be established on antibody binding cryogel affinity matrices.
|
|
| 4. |
- Ahlqvist, Josefin, et al.
(author)
-
DNA digestion and formation of DNA-network structures with Holliday junction-resolving enzyme Hjc_15-6 in conjunction with polymerase reactions
-
In: Journal of Biotechnology. - 1873-4863.
-
Journal article (peer-reviewed)abstract
- The recently identified novel Holliday junction-resolving enzyme, termed Hjc_15-6, activity investigation results imply DNA cleavage by Hjc_15-6 in a manner that potentially enhances the molecular self-assembly that may be exploited for creating DNA-networks and nanostructures. The study also demonstrates Pwo DNA polymerase acting in combination with Hjc_15-6 capability to produce large amounts of DNA that transforms into large DNA-network structures even without DNA template and primers. Furthermore, it is demonstrated that Hjc_15-6 prefers Holliday junction oligonucleotides as compared to Y-shaped oligonucleotides as well as efficiently cleaves typical branched products from isothermal DNA amplification of both linear and circular DNA templates amplified by phi29-like DNA polymerase. The assembly of large DNA network structures was observed in real time, by transmission electron microscopy, on negative stained grids that were freshly prepared, and also on the same grids after incubation for 4 days under constant cooling. Hence, Hjc_15-6 is a promising molecular tool for efficient production of various DNA origamis that may be implemented for a wide range of applications such as within medical biomaterials, catalytic materials, molecular devices and biosensors.
|
|
| 5. |
- Andersson, Henrik, et al.
(author)
-
Assaying cardiac biomarkers for toxicity testing using biosensing and cardiomyocytes derived from human embryonic stem cells
- 2010
-
In: JOURNAL OF BIOTECHNOLOGY. - : Elsevier Science B.V., Amsterdam.. - 0168-1656 .- 1873-4863. ; 150:1, s. 175-181
-
Journal article (peer-reviewed)abstract
- Human embryonic stem cell (hESC) derived cardiomyocytes are in the present study being used for testing drug-induced cardiotoxicity in a biosensor set-up. The design of an in vitro testing alternative provides a novel opportunity to surpass previous methods based on rodent cells or cell lines due to its significantly higher toxicological relevance. In this report we demonstrate how hESC-derived cardiomyocytes release detectable levels of two clinically decisive cardiac biomarkers, cardiac troponin T and fatty acid binding protein 3, when the cardiac cells are exposed to the well-known cardioactive drug compound. doxorubicin. The release is monitored by the immuno-biosensor technique surface plasmon resonance, particularly appropriate due to its capacity for parallel and high-throughput analysis in complex media.
|
|
| 6. |
- Andersson, Louise, et al.
(author)
-
Draft genome sequence of the sugar beet pathogen Rhizoctonia solani AG2-2IIIB strain BBA69670
- 2016
-
In: Journal of Biotechnology. - : Elsevier BV. - 0168-1656 .- 1873-4863. ; 222, s. 11-12
-
Journal article (peer-reviewed)abstract
- Rhizoctonia solani is a widespread plant pathogenic fungus featuring a broad host range including several economically important crops. Accordingly, genome analyses of R. solani isolates are important to uncover their pathogenic potential. Draft genome sequences for four R. solani isolates representing three of the 14 R. solani anastomosis groups (AGs) are available. Here, we present the first draft genome sequence for an R. solani AG2-2IIIB isolate that is pathogenic on sugar beet. The fungal genome was assembled in 2065 scaffolds consisting of 5826 contigs amounting to a size of about 52 Mb which is larger than any other R. solani isolate known today. Genes potentially encoding cellulolytic, lignolytic and pectinolytic enzymes were identified. (c) 2016 Elsevier B.V. All rights reserved.
|
|
| 7. |
|
|
| 8. |
- Andersson, Mariette, et al.
(author)
-
Targeted gene suppression by RNA interference: An efficient method for production of high-amylose potato lines
- 2006
-
In: Journal of Biotechnology. - : Elsevier BV. - 1873-4863 .- 0168-1656. ; 123:2, s. 137-148
-
Journal article (peer-reviewed)abstract
- Production of high-amylose potato lines can be achieved by inhibition of two genes coding for starch branching enzymes. The use of antisense technology for gene inhibition have yielded a low frequency of high-amylose lines that mostly was correlated with high numbers of integrated T-DNA copies. To investigate whether the production of high-amylose lines could be improved, RNA interference was used for gene inhibition of the genes Sbe1 and Sbe2. Two constructs with 100 bp segments (pHAS2) or 200 bp segments (pHAS3) of both branching enzyme genes were cloned as inverted repeats controlled by a potato granule-bound starch synthase promoter. The construct pHAS3 was shown to be very efficient, yielding high-amylose quality in more than 50% of the transgenic lines. An antisense construct, included in the study as a comparator, resulted in only 3% of the transgenic lines being of high-amylose type. Noticeable was also that pHAS3 yielded low T-DNA copy inserts with an average of 83% of backbone-free transgenic lines being single copy events.
|
|
| 9. |
- Andrade-Pavón, Dulce, et al.
(author)
-
Inhibition of recombinant enzyme 3-hydroxy-3-methylglutaryl-CoA reductase from Candida glabrata by α-asarone-based synthetic compounds as antifungal agents
- 2019
-
In: Journal of Biotechnology. - : Elsevier BV. - 0168-1656 .- 1873-4863. ; 292, s. 64-67
-
Journal article (peer-reviewed)abstract
- Due to increasing resistance of Candida species to antifungal drugs, especially azoles, new drugs are needed. The proposed compounds 3 and 4 are analogous to α-asarone (2), a naturally occurring potent inhibitor of HMGR with hypolipidemic and antifungal activity. We used the recombinant enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase of Candida glabrata (CgHMGR) as a model to test the effectiveness of the test compounds. Compounds 3 and 4 demonstrated inhibitory kinetics, having lower IC 50 values (42.65 μM and 28.77 μM, respectively) than compound 2 (>100 μM). The docking studies showed better binding energies for compounds 3 and 4 (−5.35 and −6.1 kcal/mol, respectively) than for compound 2 (−4.53 kcal/mol). These findings suggest that the tested compounds are better than their natural analogue. Plaque assays were performed on the C. glabrata strain CBS138 by applying ergosterol or cholesterol to evaluate the possible reversal of the inhibition induced by compounds 2, 3 and 4. Inhibition was easily suppressed in all three cases, recovering the viability of C. glabrata. These results reveal that the CgHMGR model is excellent for testing antifungals. Compound 4 produced the best effect and is herein proposed as a new potent antifungal agent.
|
|
| 10. |
- Bachinger, T., et al.
(author)
-
Monitoring cellular state transitions in a production-scale CHO-cell process using an electronic nose
- 2000
-
In: Journal of Biotechnology. - 0168-1656 .- 1873-4863. ; 76:1, s. 61-71
-
Journal article (peer-reviewed)abstract
- An electronic nose is used to monitor the bioreactor off-gas composition in perfused cultivations of a CHO-cell line producing recombinant human blood coagulation factor VIII. The applicability of the electronic nose for monitoring cellular state transitions and process control is explained. It is shown that the instrument can reveal characteristic process states related to product and lactate formation, and detect microbial infections in a very early stage of the infection. The visualization of ideal process conditions is realized by using principal component analysis (PCA) and the on-line applicability of this method is outlined. The results illustrate the potential of the electronic nose as on-line sensor for ensuring product and process quality in production-scale bioprocesses. Copyright (C) 2000 Elsevier Science B.V.An electronic nose is used to monitor the bioreactor off-gas composition in perfused cultivations of a CHO-cell line producing recombinant human blood coagulation factor VIII. The applicability of the electronic nose for monitoring cellular state transitions and process control is explained. It is shown that the instrument can reveal characteristic process states related to product and lactate formation, and detect microbial infections in a very early stage of the infection. The visualization of ideal process conditions is realized by using principal component analysis (PCA) and the on-line applicability of this method is outlined. The results illustrate the potential of the electronic nose as on-line sensor for ensuring product and process quality in production-scale bioprocesses.
|
|
