| 1. |
- Christakopoulos, Paul, et al.
(författare)
-
Functional characterization of a cellulose binding xylanase from Fusarium oxysporum
- 1996
-
Ingår i: Biotechnology letters. - 0141-5492 .- 1573-6776. ; 18:3, s. 349-354
-
Tidskriftsartikel (refereegranskat)abstract
- An extracellular endoxylanase from Fusarium oxysporum binds onto crystalline cellulose. A small peptide (~ 2kDa) could be isolated after partial proteolysis of the native protein. It consists of 18 amino acids, is located in the C-terminal region of the protein and corresponds functionally to a cellulose binding domain (CBD), the first one to be reported in a fungal xylanase. The amino acid sequence of this peptide shows no homology with any known CBD
|
|
| 2. |
- Christakopoulos, Paul, et al.
(författare)
-
Production and partial characterization of xylanase from Fusarium oxysporum
- 1996
-
Ingår i: Bioresource Technology. - 0960-8524 .- 1873-2976. ; 58:2, s. 115-119
-
Tidskriftsartikel (refereegranskat)abstract
- Production of xylanase by Fusarium oxysporum strain F3 was enhanced by optimization of initial pH of the culture medium, the type and concentration of nitrogen and carbon source, and the growth temperature. Under these conditions, yields as high as 245 U/ml of culture medium were obtained. The most important characteristic of the enzyme is its high pH stability. It retained 80 and 66% of the activity at pH 9.0 after 24 h at 4 and 30°C, respectively. Chromogenic (fluorogenic) 4-methylumbelliferyl-β-glycosides of xylose (MUX) and xylobiose (MUX2) were used to characterize xylanase multienzyme components, after separation by isoelectric focusing. The zymogram indicated one major, one minor xylanase and one active β-xylosidase exhibiting pI values of 9.5, 6.5 and 3.8, respectively
|
|
| 3. |
- Christakopoulos, Paul, et al.
(författare)
-
Purification and characterization of two low molecular mass alkaline xylanases from Fusarium oxysporum F3
- 1996
-
Ingår i: Journal of Biotechnology. - : Elsevier BV. - 0168-1656 .- 1873-4863. ; 51:2, s. 181-189
-
Tidskriftsartikel (refereegranskat)abstract
- Two low molecular mass endo-1,4-β-d-xylanases from Fusarium oxysporum were purified to homogeneity by gel-filtration and ion-exchange chromatography. They exhibit molecular masses of 20.8 (xylanase I) and 23.5 (xylanase II) kDa, and isoelectric points of 9.5 and 8.45–8.70, respectively. Both xylanases display remarkable pH (9.0) stability. At 40 to 55 °C xylanase II is more thermostable than xylanase I but less active on xylan. In contrast to xylanase I, xylanase II is able to hydrolyze 1-O-4-methylumbelliferyl-(β-d-glucopyranosyl)-β-d-xylopyranoside (muxg). Neither of these enzymes hydrolyze xylotriose. They bind on crystalline cellulose but not on insoluble xylan. Analysis of reaction mixtures by high pressure liquid chromatography revealed that both enzymes cleave preferentially the internal glycosidic bonds of xylopentaose and oat spelts xylan. Thus the purified enzymes appeared to be true endo-β-1,4-xylanases. The amino terminal sequences of xylanases I and II show no homology. Xylanase I shows high similarity with alkaline low molecular mass xylanases of family G/11.
|
|
| 4. |
- Zou, JY, et al.
(författare)
-
Crystallographic evidence for substrate ring distortion and protein conformational changes during catalysis in cellobiohydrolase Cel6A from Trichoderma reesei
- 1999
-
Ingår i: STRUCTURE WITH FOLDING & DESIGN. - 0969-2126. ; 7:9, s. 1035-1045
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Cel6A is one of the two cellobiohydrolases produced by Trichoderma reesei. The catalytic core has a structure that is a variation of the classic TIM barrel. The active site is located inside a tunnel, the roof of which is formed mainly by a pair of loops. RESULTS: We describe three new ligand complexes. One is the structure of the wild-type enzyme in complex with a nonhydrolysable cello-oligosaccharide, methyl 4-S-beta-cellobiosyl-4-thio-beta-cellobioside (Glc)(2)-S-(Glc)(2), which differs from a cellotetraose in the nature of the central glycosidic linkage where a sulphur atom replaces an oxygen atom. The second structure is a mutant, Y169F, in complex with the same ligand, and the third is the wild-type enzyme in complex with m-iodobenzyl beta-D-glucopyranosyl-beta(1,4)-D-xylopyranoside (IBXG). CONCLUSIONS: The (Glc)(2)-S-(Glc)(2) ligand binds in the -2 to +2 sites in both the wild-type and mutant enzymes. The glucosyl unit in the -1 site is distorted from the usual chair conformation in both structures. The IBXG ligand binds in the -2 to +1 sites, with the xylosyl unit in the -1 site where it adopts the energetically favourable chair conformation. The -1 site glucosyl of the (Glc)(2)-S-(Glc)(2) ligand is unable to take on this conformation because of steric clashes with the protein. The crystallographic results show that one of the tunnel-forming loops in Cel6A is sensitive to modifications at the active site, and is able to take on a number of different conformations. One of the conformational changes disrupts a set of interactions at the active site that we propose is an integral part of the reaction mechanism.
|
|
