| 2. |
- Jong, Wouter S. P., et al.
(författare)
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Application of an E. coli signal sequence as a versatile inclusion body tag
- 2017
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Ingår i: Microbial Cell Factories. - : Springer Science and Business Media LLC. - 1475-2859. ; 16
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Tidskriftsartikel (refereegranskat)abstract
- Background: Heterologous protein production in Escherichia coli often suffers from bottlenecks such as proteolytic degradation, complex purification procedures and toxicity towards the expression host. Production of proteins in an insoluble form in inclusion bodies (IBs) can alleviate these problems. Unfortunately, the propensity of heterologous proteins to form IBs is variable and difficult to predict. Hence, fusing the target protein to an aggregation prone polypeptide or IB-tag is a useful strategy to produce difficult-to-express proteins in an insoluble form. Results: When screening for signal sequences that mediate optimal targeting of heterologous proteins to the periplasmic space of E. coli, we observed that fusion to the 39 amino acid signal sequence of E. coli TorA (ssTorA) did not promote targeting but rather directed high-level expression of the human proteins hEGF, Pla2 and IL-3 in IBs. Further analysis revealed that ssTorA even mediated IB formation of the highly soluble endogenous E. coli proteins TrxA and MBP. The ssTorA also induced aggregation when fused to the C-terminus of target proteins and appeared functional as IB-tag in E. coli K-12 as well as B strains. An additive effect on IB-formation was observed upon fusion of multiple ssTorA sequences in tandem, provoking almost complete aggregation of TrxA and MBP. The ssTorA-moiety was successfully used to produce the intrinsically unstable hEGF and the toxic fusion partner SymE, demonstrating its applicability as an IB-tag for difficult-to-express and toxic proteins. Conclusions: We present proof-of-concept for the use of ssTorA as a small, versatile tag for robust E. coli-based expression of heterologous proteins in IBs.
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| 3. |
- Mills, Gina, 1959, et al.
(författare)
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Ozone pollution will compromise efforts to increase global wheat production
- 2018
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Ingår i: Global Change Biology. - : Wiley. - 1354-1013 .- 1365-2486. ; 24:8, s. 3560-3574
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Tidskriftsartikel (refereegranskat)abstract
- Introduction of high-performing crop cultivars and crop/soil water management practices that increase the stomatal uptake of carbon dioxide and photosynthesis will be instrumental in realizing the United Nations Sustainable Development Goal (SDG) of achieving food security. To date, however, global assessments of how to increase crop yield have failed to consider the negative effects of tropospheric ozone, a gaseous pollutant that enters the leaf stomatal pores of plants along with carbon dioxide, and is increasing in concentration globally, particularly in rapidly developing countries. Earlier studies have simply estimated that the largest effects are in the areas with the highest ozone concentrations. Using a modelling method that accounts for the effects of soil moisture deficit and meteorological factors on the stomatal uptake of ozone, we show for the first time that ozone impacts on wheat yield are particularly large in humid rain-fed and irrigated areas of major wheat-producing countries (e.g. United States, France, India, China and Russia). Averaged over 2010-2012, we estimate that ozone reduces wheat yields by a mean 9.9% in the northern hemisphere and 6.2% in the southern hemisphere, corresponding to some 85 Tg (million tonnes) of lost grain. Total production losses in developing countries receiving Official Development Assistance are 50% higher than those in developed countries, potentially reducing the possibility of achieving UN SDG2. Crucially, our analysis shows that ozone could reduce the potential yield benefits of increasing irrigation usage in response to climate change because added irrigation increases the uptake and subsequent negative effects of the pollutant. We show that mitigation of air pollution in a changing climate could play a vital role in achieving the above-mentioned UN SDG, while also contributing to other SDGs related to human health and well-being, ecosystems and climate change.
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