| 1. |
- Vardakou, Maria, et al.
(författare)
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A family 10 Thermoascus aurantiacus xylanase utilizes arabinose decorations of xylan as significant substrate specificity determinants
- 2005
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 352:5, s. 1060-1067
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Tidskriftsartikel (refereegranskat)abstract
- Xylan, which is a key component of the plant cell wall, consists of a backbone of β-1,4-linked xylose residues that are decorated with arabinofuranose, acetyl, 4-O-methyl d-glucuronic acid and ferulate. The backbone of xylan is hydrolysed by endo-β1,4-xylanases (xylanases); however, it is unclear whether the various side-chains of the polysaccharide are utilized by these enzymes as significant substrate specificity determinants. To address this question we have determined the crystal structure of a family 10 xylanase from Thermoascus aurantiacus, in complex with xylobiose containing an arabinofuranosyl-ferulate side-chain. We show that the distal glycone subsite of the enzyme makes extensive direct and indirect interactions with the arabinose side-chain, while the ferulate moiety is solvent-exposed. Consistent with the 3D structural data, the xylanase displays fourfold more activity against xylotriose in which the non-reducing moiety is linked to an arabinose side-chain, compared to the undecorated form of the oligosacchairde. These data indicate that the sugar decorations of xylans in the T. aurantiacus family 10 xylanase, rather than simply being accommodated, can be significant substrate specificity determinants.
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| 2. |
- Vardakou, Maria, et al.
(författare)
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Understanding the structural basis for substrate and inhibitor recognition in Eukaryotic GH11 Xylanases
- 2008
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 375:5, s. 1293-1305
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Tidskriftsartikel (refereegranskat)abstract
- Endo-β1,4-xylanases (xylanases) hydrolyse the β1,4 glycosidic bonds in the backbone of xylan. Although xylanases from glycoside hydrolase family 11 (GH11) have been extensively studied, several issues remain unresolved. Thus, the mechanism by which these enzymes hydrolyse decorated xylans is unclear and the structural basis for the variation in catalytic activity within this family is unknown. Furthermore, the mechanism for the differences in the inhibition of fungal GH11 enzymes by the wheat protein XIP-I remains opaque. To address these issues we report the crystal structure and biochemical properties of the Neocallimastix patriciarum xylanase NpXyn11A, which displays unusually high catalytic activity and is one of the few fungal GH11 proteins not inhibited by XIP-I. Although the structure of NpXyn11A could not be determined in complex with substrates, we have been able to investigate how GH11 enzymes hydrolyse decorated substrates by solving the crystal structure of a second GH11 xylanase, EnXyn11A (encoded by an environmental DNA sample), bound to ferulic acid-1,5-arabinofuranose-α1,3-xylotriose (FAX3). The crystal structure of the EnXyn11A–FAX3 complex shows that solvent exposure of the backbone xylose O2 and O3 groups at subsites −3 and +2 allow accommodation of α1,2-linked 4-methyl-D-glucuronic acid and L-arabinofuranose side chains. Furthermore, the ferulated arabinofuranose side chain makes hydrogen bonds and hydrophobic interactions at the +2 subsite, indicating that the decoration may represent a specificity determinant at this aglycone subsite. The structure of NpXyn11A reveals potential −3 and +3 subsites that are kinetically significant. The extended substrate-binding cleft of NpXyn11A, compared to other GH11 xylanases, may explain why the Neocallimastix enzyme displays unusually high catalytic activity. Finally, the crystal structure of NpXyn11A shows that the resistance of the enzyme to XIP-I is not due solely to insertions in the loop connecting β strands 11 and 12, as suggested previously, but is highly complex.
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| 3. |
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| 4. |
- Daniel, Terry C., et al.
(författare)
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Contributions of cultural services to the ecosystem services agenda
- 2012
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 109:23, s. 8812-8819
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Tidskriftsartikel (refereegranskat)abstract
- Cultural ecosystem services (ES) are consistently recognized but not yet adequately defined or integrated within the ES framework. A substantial body of models, methods, and data relevant to cultural services has been developed within the social and behavioral sciences before and outside of the ES approach. A selective review of work in landscape aesthetics, cultural heritage, outdoor recreation, and spiritual significance demonstrates opportunities for operationally defining cultural services in terms of socioecological models, consistent with the larger set of ES. Such models explicitly link ecological structures and functions with cultural values and benefits, facilitating communication between scientists and stakeholders and enabling economic, multicriterion, deliberative evaluation and other methods that can clarify tradeoffs and synergies involving cultural ES. Based on this approach, a common representation is offered that frames cultural services, along with all ES, by the relative contribution of relevant ecological structures and functions and by applicable social evaluation approaches. This perspective provides a foundation for merging ecological and social science epistemologies to define and integrate cultural services better within the broader ES framework.
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| 5. |
- de Gonzalez, Amy Berrington, et al.
(författare)
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Body-Mass Index and Mortality among 1.46 Million White Adults.
- 2010
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Ingår i: New England Journal of Medicine. - : MASSACHUSETTS MEDICAL SOC. - 0028-4793 .- 1533-4406. ; 363:23, s. 2211-2219
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Tidskriftsartikel (refereegranskat)abstract
- Background: A high body-mass index (BMI, the weight in kilograms divided by the square of the height in meters) is associated with increased mortality from cardiovascular disease and certain cancers, but the precise relationship between BMI and all-cause mortality remains uncertain. Methods: We used Cox regression to estimate hazard ratios and 95% confidence intervals for an association between BMI and all-cause mortality, adjusting for age, study, physical activity, alcohol consumption, education, and marital status in pooled data from 19 prospective studies encompassing 1.46 million white adults, 19 to 84 years of age (median, 58). Results: The median baseline BMI was 26.2. During a median follow-up period of 10 years (range, 5 to 28), 160,087 deaths were identified. Among healthy participants who never smoked, there was a J-shaped relationship between BMI and all-cause mortality. With a BMI of 22.5 to 24.9 as the reference category, hazard ratios among women were 1.47 (95 percent confidence interval [CI], 1.33 to 1.62) for a BMI of 15.0 to 18.4; 1.14 (95% CI, 1.07 to 1.22) for a BMI of 18.5 to 19.9; 1.00 (95% CI, 0.96 to 1.04) for a BMI of 20.0 to 22.4; 1.13 (95% CI, 1.09 to 1.17) for a BMI of 25.0 to 29.9; 1.44 (95% CI, 1.38 to 1.50) for a BMI of 30.0 to 34.9; 1.88 (95% CI, 1.77 to 2.00) for a BMI of 35.0 to 39.9; and 2.51 (95% CI, 2.30 to 2.73) for a BMI of 40.0 to 49.9. In general, the hazard ratios for the men were similar. Hazard ratios for a BMI below 20.0 were attenuated with longer-term follow-up. Conclusions: In white adults, overweight and obesity (and possibly underweight) are associated with increased all-cause mortality. All-cause mortality is generally lowest with a BMI of 20.0 to 24.9. N Engl J Med 2010;363:2211-9.
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| 6. |
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| 7. |
- Hoy, Damian, et al.
(författare)
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Lessons learnt from a three-year pilot field epidemiology training programme
- 2017
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Ingår i: Western Pacific surveillance and response journal : WPSAR. - : World Health Organization, Western Pacific Regional Office. - 2094-7313 .- 2094-7321. ; 8:3, s. 21-26
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Tidskriftsartikel (refereegranskat)abstract
- PROBLEM: The Pacific region has widely dispersed populations, limited financial and human resources and a high burden of disease. There is an urgent need to improve the availability, reliability and timeliness of useable health data.CONTEXT: The purpose of this paper is to share lessons learnt from a three-year pilot field epidemiology training programme that was designed to respond to these Pacific health challenges. The pilot programme built on and further developed an existing field epidemiology training programme for Pacific health staff.ACTION: The programme was delivered in country by epidemiologists working for Pacific Public Health Surveillance Network partners. The programme consisted of five courses: four one-week classroom-based courses and one field epidemiology project. Sessions were structured so that theoretical understanding was achieved through interaction and reinforced through practical hands-on group activities, case studies and other interactive practical learning methods.OUTCOME: As of September 2016, 258 students had commenced the programme. Twenty-six course workshops were delivered and one cohort of students had completed the full five-course programme. The programme proved popular and gained a high level of student engagement.DISCUSSION: Face-to-face delivery, a low student-to-facilitator ratio, substantial group work and practical exercises were identified as key factors that contributed to the students developing skills and confidence. Close engagement of leaders and the need to quickly evaluate and adapt the curriculum were important lessons, and the collaboration between external partners was considered important for promoting a harmonized approach to health needs in the Pacific.
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| 8. |
- Montanier, Cedric, et al.
(författare)
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The Active Site of a Carbohydrate Esterase Displays Divergent Catalytic and Noncatalytic Binding Functions
- 2009
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Ingår i: PLoS Biology. - : Public Library of Science (PLoS). - 1545-7885. ; 7:3, s. 687-697
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Tidskriftsartikel (refereegranskat)abstract
- Multifunctional proteins, which play a critical role in many biological processes, have typically evolved through the recruitment of different domains that have the required functional diversity. Thus the different activities displayed by these proteins are mediated by spatially distinct domains, consistent with the specific chemical requirements of each activity. Indeed, current evolutionary theory argues that the colocalization of diverse activities within an enzyme is likely to be a rare event, because it would compromise the existing activity of the protein. In contrast to this view, a potential example of multifunctional recruitment into a single protein domain is provided by CtCel5C-CE2, which contains an N-terminal module that displays cellulase activity and a C-terminal module, CtCE2, which exhibits a noncatalytic cellulose-binding function but also shares sequence identity with the CE2 family of esterases. Here we show that, unlike other CE2 members, the CtCE2 domain displays divergent catalytic esterase and noncatalytic carbohydrate binding functions. Intriguingly, these diverse activities are housed within the same site on the protein. Thus, a critical component of the active site of CtCE2, the catalytic Ser-His dyad, in harness with inserted aromatic residues, confers noncatalytic binding to cellulose whilst the active site of the domain retains its esterase activity. CtCE2 catalyses deacetylation of noncellulosic plant structural polysaccharides to deprotect these substrates for attack by other enzymes. Yet it also acts as a cellulose-binding domain, which promotes the activity of the appended cellulase on recalcitrant substrates. The CE2 family encapsulates the requirement for multiple activities by biocatalysts that attack challenging macromolecular substrates, including the grafting of a second, powerful and discrete noncatalytic binding functionality into the active site of an enzyme. This article provides a rare example of "gene sharing,'' where the introduction of a second functionality into the active site of an enzyme does not compromise the original activity of the biocatalyst.
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| 9. |
- Montanier, Cedric Y., et al.
(författare)
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A novel, noncatalytic carbohydrate-binding module displays specificity for galactose-containing polysaccharides through calcium-mediated oligomerization
- 2011
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Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 286:25
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Tidskriftsartikel (refereegranskat)abstract
- The enzymic degradation of plant cell walls plays a central role in the carbon cycle and is of increasing environmental and industrial significance. The catalytic modules of enzymes that catalyze this process are generally appended to noncatalytic carbohydrate-binding modules (CBMs). CBMs potentiate the rate of catalysis by bringing their cognate enzymes into intimate contact with the target substrate. A powerful plant cell wall-degrading system is the Clostridium thermocellum multienzyme complex, termed the "cellulosome." Here, we identify a novel CBM (CtCBM62) within the large C. thermocellum cellulosomal protein Cthe_2193 (defined as CtXyl5A), which establishes a new CBM family. Phylogenetic analysis of CBM62 members indicates that a circular permutation occurred within the family. CtCBM62 binds to D-galactose and L-arabinopyranose in either anomeric configuration. The crystal structures of CtCBM62, in complex with oligosaccharides containing alpha- and beta-galactose residues, show that the ligand-binding site in the beta-sandwich protein is located in the loops that connect the two beta-sheets. Specificity is conferred through numerous interactions with the axial O4 of the target sugars, a feature that distinguishes galactose and arabinose from the other major sugars located in plant cell walls. CtCBM62 displays tighter affinity for multivalent ligands compared with molecules containing single galactose residues, which is associated with precipitation of these complex carbohydrates. These avidity effects, which confer the targeting of polysaccharides, are mediated by calcium-dependent oligomerization of the CBM.
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| 10. |
- von Schantz, Laura, et al.
(författare)
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Affinity maturation generates greatly improved xyloglucan-specific carbohydrate binding modules
- 2009
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Ingår i: BMC Biotechnology. - : Springer Science and Business Media LLC. - 1472-6750. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Background: Molecular evolution of carbohydrate binding modules (CBM) is a new approach for the generation of glycan-specific molecular probes. To date, the possibility of performing affinity maturation on CBM has not been investigated. In this study we show that binding characteristics such as affinity can be improved for CBM generated from the CBM4-2 scaffold by using random mutagenesis in combination with phage display technology. Results: Two modified proteins with greatly improved affinity for xyloglucan, a key polysaccharide abundant in the plant kingdom crucial for providing plant support, were generated. Both improved modules differ from other existing xyloglucan probes by binding to galactose-decorated subunits of xyloglucan. The usefulness of the evolved binders was verified by staining of plant sections, where they performed better than the xyloglucan-binding module from which they had been derived. They discriminated non-fucosylated from fucosylated xyloglucan as shown by their ability to stain only the endosperm, rich in non-fucosylated xyloglucan, but not the integument rich in fucosylated xyloglucan, on tamarind seed sections. Conclusion: We conclude that affinity maturation of CBM selected from molecular libraries based on the CBM4-2 scaffold is possible and has the potential to generate new analytical tools for detection of plant carbohydrates.
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