| 1. |
- Gaber, Alexander, et al.
(author)
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High expression of tumour-associated trypsin inhibitor correlates with liver metastasis and poor prognosis in colorectal cancer
- 2009
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In: British Journal of Cancer. - : Springer Science and Business Media LLC. - 0007-0920 .- 1532-1827. ; 100:10, s. 1540-1548
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Journal article (peer-reviewed)abstract
- Increased expression of tumour-associated trypsin inhibitor (TATI) in tumour tissue and/or serum has been associated with poor survival in various cancer forms. Moreover, a proinvasive function of TATI has been shown in colon cancer cell lines. In this study, we have examined the prognostic significance of tumour-specific TATI expression in colorectal cancer, assessed by immunohistochemistry (IHC) on tissue microarrays (TMAs) with tumour specimens from two independent patient cohorts. Kaplan-Meier analysis and Cox proportional hazards modelling were used to estimate time to recurrence, disease-free survival and overall survival. In both cohorts, a high (>50% of tumour cells) TATI expression was an independent predictor of a significantly shorter overall survival. In cohort II, in multivariate analysis including age, gender, disease stage, differentiation grade, vascular invasion and carcinoembryonal antigen (CEA), high TATI expression was associated with a significantly decreased overall survival (HR=1.82; 95% CI=1.19-2.79) and disease-free survival (HR=1.56; 95% CI=1.05-2.32) in curatively treated patients. Moreover, there was an increased risk for liver metastasis in both cohorts that remained significant in multivariate analysis in cohort II (HR=2.85; 95% CI=1.43-5.66). In conclusion, high TATI expression is associated with liver metastasis and is an independent predictor of poor prognosis in patients with colorectal cancer.
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| 2. |
- Birgisson, Hakon, et al.
(author)
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A new thermostable alpha-L-arabinofuranosidase from a novel thermophilic bacterium
- 2004
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In: Biotechnology Letters. - 1573-6776. ; 26:17, s. 1347-1351
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Journal article (peer-reviewed)abstract
- An alpha-L-arabinofuranosidase gene was identified in a sequenced genome of a novel thermophilic bacterium, which belongs to the recently described phylum of Thermomicrobia. Amino acid sequence comparison of the enzyme (designated AraF) revealed similarity to glycoside hydrolases of family 51. The gene was cloned into Escherichia coli and its recombinant product expressed and purified. The enzyme appeared to be a hexamer. AraF was optimally active at 70degreesC (over 10 min) and pH 6 having 92% residual activity after 1 h at 70degreesC. AraF had a K-m value of 0.6 rum and V-max value of 122 U mg(-1) on p-nitrophenyl-alpha-L-arabinofuranoside. AraF was almost equally active on branched arabinan and debranched arabinan, properties not previously found in alpha-L-arabinofuranosidases in GH family 51.
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| 3. |
- Birgisson, Hakon, et al.
(author)
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Cold adapted yeasts as producers of cold active polygalacturonases
- 2003
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In: Extremophiles. - : Springer Science and Business Media LLC. - 1433-4909 .- 1431-0651. ; 7:3, s. 185-193
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Journal article (peer-reviewed)abstract
- Eight cold-adapted, polygalacturonase-producing yeasts belonging to four species were isolated from frozen environmental samples in Iceland. They were identified as Cystofilobasidium lari-marini, Cystofilobasidium capitatum, Cryptococcus macerans and Cryptococcus aquaticus species by sequence analysis of rDNA regions. Growth behavior of the isolates was investigated. All strains could grow at 2°C. Addition of glucose to pectin-containing culture medium had a repressive effect on enzyme production except for C. aquaticus, which showed increased polygalacturonase activity. Optimal temperature for enzyme production for the Cystofilobasidium strains was 14°C, while that for the Cryptococcus strains was lower. Among the isolates, C. lari-marini S3B produced highest levels of enzyme activity at pH 3.2. Preliminary characterization of the polygalacturonases in the culture supernatant showed the enzyme from Cystofilobasidium strains to be optimally active at 40°C and pH 5, and that from the Cryptococcus strains at 50°C and pH 4. The polygalacturonase from C. macerans started to lose activity after 1 h of incubation at 40°C, while that from the other strains had already lost activity at 30°C. All the strains except C. aquaticus produced isoenzymes of polyglacturonase. In addition to polygalacturonase, the Cystofilobasidium strains produced pectin lyase, C. aquaticus pectin esterase, and C. macerans pectin lyase, pectate lyase and pectin esterase.
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| 4. |
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| 5. |
- Birgisson, Hakon
(author)
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Glycoside hydrolases from extremophiles isolated in Iceland. alpha-L-Rhamnosidases and pectin degrading enzymes
- 2004
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Doctoral thesis (other academic/artistic)abstract
- Extreme environments (high and low temperature, high and low pH, etc.) are inhabited by a vast and diverse flora of microorganisms, which are adapted to these environments (extremophiles). The enzymes produced by extremophiles are active under these harsh conditions and are thus applicable in processes, where conditions are outside the working range of enzymes originating from mesophiles. In this thesis, both cold-adapted and heat-adapted glycoside hydrolases identified in microorganisms isolated in cold and hot environments in Iceland are described. Screening of soil samples for pectinolytic microorganisms, resulted in isolation of four cold-adapted yeast species, which all excreted one or more polygalacturonases. Analysis of the 5.8S rDNA and the 26S rDNA D1/D2 domain, revealed that the yeasts were most closely related to Cystofilobasidium capitatum, Cystofilobasidium lari-marini, Cryptococcus macerans and Cryptococcus aquaticus. Examination of the polygalacturonase activities revealed that at 2 °C, all samples showed 8-18% activity of their activities at optimal temperatures. Screening of a sequenced genome of a novel thermophilic bacterium resulted in isolation, cloning and expression of three genes. Two of the genes encoded for alpha-L-rhamnosidases and one for alpha-L-arabinofuranosidase. All purified recombinant products were found to have broad pH activity curves, temperature optima at 70 °C and relatively high thermostability. One of the recombinant alpha-L-rhamnosidases was produced with a His tag, to facilitate purification of large quantities of the enzyme. The purified enzyme was subsequently used for crystallization. Two crystal forms were obtained. X-ray analysis indicated that one of the crystal forms may be useful for structural determination of the enzyme. Finally, the recombinant cells producing one of the two recombinant alpha-L-rhamnosidases were immobilized in alginate beads, which were used for construction of a bioreactor for production of L-rhamnose from a natural substrate at elevated temperatures.
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| 6. |
- Birgisson, Hakon, et al.
(author)
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Immobilization of a recombinant Escherichia coli producing a thermostable alpha-L-rhamnosidase: Creation of a bioreactor for hydrolyses of naringin
- 2007
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In: Enzyme and Microbial Technology. - : Elsevier BV. - 0141-0229. ; 40:5, s. 1181-1187
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Journal article (peer-reviewed)abstract
- An U-L-rhamnosidase (E.C. 3.2.1.40) from a newly discovered thermophilic bacterium was expressed in Escherichia coli BL21 DE3 pRIL cells. The cells were immobilized in Ca2+-alginate beads. The temperature of 50 degrees C used in reactions, appeared to be sufficient for making the mesophilic strain porous enough for the substrate to access the cloned thermostable enzyme. Pretreatment of cells with heat or lysozyme prior to bead formation did not improve the results. The best cell concentration (w/w) for bead preparation was found to be 0.0 192 g ml(-1) and stability of beads increased if CaCl2 concentration in buffers and substrate was kept at 50 mM. In a 60 min assay, the optimal pH of the entrapped cells was found to be 7.8 and the optimal temperature 60 degrees C. By packing the beads in a column, a bioreactor for production Of L-rhamnose from naringin was created. Full degradation of 7.9 mM naringin could be reached by running the reactor at 1 ml min(-1) at 50 degrees C. The optimal running temperature of the reactor was found to be 50 degrees C and the reactor was fully stable over 3 days at that temperature. On the fourth day, substrate degradation capacity had decreased by 10-15%. (c) 2006 Elsevier Inc. All rights reserved.
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| 7. |
- Birgisson, Hakon, et al.
(author)
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Two new thermostable alpha-L-rhamnosidases from a novel thermophilic bacterium
- 2004
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In: Enzyme and Microbial Technology. - : Elsevier BV. - 0141-0229. ; 34:6, s. 561-571
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Journal article (peer-reviewed)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.
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| 8. |
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