Ultraviolet absorbance of Sphagnum magellanicum, S. fallax and S. fuscum extracts with seasonal and species-specific variation.

Bryophytes, including Sphagnum, are common species in alpine and boreal regions especially on mires, where full sunlight exposes the plants to the damaging effects of UV radiation. Sphagnum species containing UV-protecting compounds might offer a biomass source for nature-based sunscreens to replace the synthetic ones. In this study, potential compounds and those linked in cell wall structures were obtained by using methanol and alkali extractions and the UV absorption of these extracts from three common Sphagnum moss species Sphagnum magellanicum, Sphagnum fuscum and Sphagnum fallax collected in spring and autumn from western Finland are described. Absorption spectrum screening (200-900 nm) and luminescent biosensor (Escherichia coli DPD2794) methodology were used to examine and compare the protection against UV radiation. Additionally, the antioxidant potential was evaluated using hydrogen peroxide scavenging (SCAV), oxygen radical absorbance capacity (ORAC) and ferric reducing absorbance capacity (FRAP). Total phenolic content was also determined using the Folin-Ciocalteu method. The results showed that methanol extractable compounds gave higher UV absorption with the used methods. Sphagnum fallax appeared to give the highest absorption in UV-B and UV-A wavelengths. In all assays except the SCAV test, the methanol extracts of Sphagnum samples collected in autumn indicated the highest antioxidant capacity and polyphenol content. Sphagnum fuscum implied the highest antioxidant capacity and phenolic content. There was low antioxidant and UV absorption provided by the alkali extracts of these three species.

Backbone circularization of Bacillus subtilis family 11 xylanase increases its thermostability and its resistance against aggregation.

The activity of proteins is dictated by their three-dimensional structure, the native state, and is influenced by their ability to remain in or return to the folded native state under physiological conditions. Backbone circularization is thought to increase protein stability by decreasing the conformational entropy in the unfolded state. A positive effect of circularization on stability has been shown for several proteins. Here, we report the development of a cloning standard that facilitates implementing the SICLOPPS technology to circularize proteins of interest using split inteins. To exemplify the usage of the cloning standard we constructed two circularization vectors based on the Npu DnaE and gp41-1 split inteins, respectively. We use these vectors to overexpress in Escherichia coli circular forms of the Bacillus subtilis enzyme family 11 xylanase that differ in the identity and number of additional amino acids used for circularization (exteins). We found that the variant circularized with only one additional serine has increased thermostability of 7 °C compared to native xylanase. The variant circularized with six additional amino acids has only a mild increase in thermostability compared to the corresponding exteins-bearing linear xylanase, but is less stable than native xylanase. However, this circular xylanase retains more than 50% of its activity after heat shock at elevated temperatures, while native xylanase and the corresponding exteins-bearing linear xylanase are largely inactivated. We correlate this residual activity to the fewer protein aggregates found in the test tubes of circular xylanase after heat shock, suggesting that circularization protects the protein from aggregation under these conditions. Taken together, these data indicate that backbone circularization has a positive effect on xylanase and can lead to increased thermostability, provided the appropriate exteins are selected. We believe that our cloning standard and circularization vectors will facilitate testing the effects of circularization on other proteins.