| 1. |
- Wiebe, M. G., et al.
(författare)
-
A novel aldose-aldose oxidoreductase for co-production of D-xylonate and xylitol from D-xylose with Saccharomyces cerevisiae
- 2015
-
Ingår i: Applied Microbiology and Biotechnology. - : Springer Science and Business Media LLC. - 1432-0614 .- 0175-7598. ; 99:22, s. 9439-47
-
Tidskriftsartikel (refereegranskat)abstract
- An open reading frame CC1225 from the Caulobacter crescentus CB15 genome sequence belongs to the Gfo/Idh/MocA protein family and has 47 % amino acid sequence identity with the glucose-fructose oxidoreductase from Zymomonas mobilis (Zm GFOR). We expressed the ORF CC1225 in the yeast Saccharomyces cerevisiae and used a yeast strain expressing the gene coding for Zm GFOR as a reference. Cell extracts of strains overexpressing CC1225 (renamed as Cc aaor) showed some Zm GFOR type of activity, producing D-gluconate and D-sorbitol when a mixture of D-glucose and D-fructose was used as substrate. However, the activity in Cc aaor expressing strain was >100-fold lower compared to strains expressing Zm gfor. Interestingly, C. crescentus AAOR was clearly more efficient than the Zm GFOR in converting in vitro a single sugar substrate D-xylose (10 mM) to xylitol without an added cofactor, whereas this type of activity was very low with Zm GFOR. Furthermore, when cultured in the presence of D-xylose, the S. cerevisiae strain expressing Cc aaor produced nearly equal concentrations of D-xylonate and xylitol (12.5 g D-xylonate l(-1) and 11.5 g D-xylitol l(-1) from 26 g D-xylose l(-1)), whereas the control strain and strain expressing Zm gfor produced only D-xylitol (5 g l(-1)). Deletion of the gene encoding the major aldose reductase, Gre3p, did not affect xylitol production in the strain expressing Cc aaor, but decreased xylitol production in the strain expressing Zm gfor. In addition, expression of Cc aaor together with the D-xylonolactone lactonase encoding the gene xylC from C. crescentus slightly increased the final concentration and initial volumetric production rate of both D-xylonate and D-xylitol. These results suggest that C. crescentus AAOR is a novel type of oxidoreductase able to convert the single aldose substrate D-xylose to both its oxidized and reduced product.
|
|
| 2. |
|
|
| 3. |
- Borrega, M., et al.
(författare)
-
Utilizing and Valorizing Oat and Barley Straw as an Alternative Source of Lignocellulosic Fibers
- 2022
-
Ingår i: Materials. - : MDPI AG. - 1996-1944 .- 1996-1944. ; 15:21
-
Tidskriftsartikel (refereegranskat)abstract
- The transition to sustainable, biodegradable, and recyclable materials requires new sources of cellulose fibers that are already used in large volumes by forest industries. Oat and barley straws provide interesting alternatives to wood fibers in lightweight material applications because of their similar chemical composition. Here we investigate processing and material forming concepts, which would enable strong fiber network structures for various applications. The idea is to apply mild pretreatment processing that could be distributed locally so that the logistics of the raw material collection could be made efficient. The actual material production would then combine foam-forming and hot-pressing operations that allow using all fractions of fiber materials with minimal waste. We aimed to study the technical features of this type of processing on a laboratory scale. The homogeneity of the sheet samples was very much affected by whether the raw material was mechanically refined or not. Straw fibers did not form a bond spontaneously with one another after drying the sheets, but their effective bonding required a subsequent hot pressing operation. The mechanical properties of the formed materials were at a similar level as those of the conventional wood-fiber webs. In addition to the technical aspects of materials, we also discuss the business opportunities and system-level requirements of using straw as an alternative source of lignocellulosic fibers.
|
|
| 4. |
|
|
| 5. |
|
|
| 6. |
|
|
| 7. |
|
|
| 8. |
- Rudberg, P C, et al.
(författare)
-
Leukotriene A(4) hydrolase - Identification of a common carboxylate recognition site for the epoxide hydrolase and aminopeptidase substrates
- 2004
-
Ingår i: Journal of Biological Chemistry. - 1083-351X .- 0021-9258. ; 279:26, s. 27376-27382
-
Tidskriftsartikel (refereegranskat)abstract
- Leukotriene ( LT) A(4) hydrolase is a bifunctional zinc metalloenzyme, which converts LTA(4) into the neutrophil chemoattractant LTB4 and also exhibits an anion-dependent aminopeptidase activity. In the x-ray crystal structure of LTA(4) hydrolase, Arg(563) and Lys(565) are found at the entrance of the active center. Here we report that replacement of Arg(563), but not Lys(565), leads to complete abrogation of the epoxide hydrolase activity. However, mutations of Arg(563) do not seem to affect substrate binding strength, because values of K-i for LTA(4) are almost identical for wild type and ( R563K) LTA(4) hydrolase. These results are supported by the 2.3-Angstrom crystal structure of (R563A) LTA(4) hydrolase, which does not reveal structural changes that can explain the complete loss of enzyme function. For the aminopeptidase reaction, mutations of Arg(563) reduce the catalytic activity (V-max = 0.3 - 20%), whereas mutations of Lys(565) have limited effect on catalysis (V-max = 58 - 108%). However, in (K565A)- and (K565M) LTA(4) hydrolase, i.e. mutants lacking a positive charge, values of the Michaelis constant for alanine-p-nitroanilide increase significantly (K-m = 480 - 640%). Together, our data indicate that Arg(563) plays an unexpected, critical role in the epoxide hydrolase reaction, presumably in the positioning of the carboxylate tail to ensure perfect substrate alignment along the catalytic elements of the active site. In the aminopeptidase reaction, Arg(563) and Lys(565) seem to cooperate to provide sufficient binding strength and productive alignment of the substrate. In conclusion, Arg(563) and Lys(565) possess distinct roles as carboxylate recognition sites for two chemically different substrates, each of which is turned over in separate enzymatic reactions catalyzed by LTA(4) hydrolase.
|
|
