Functioning methionine sulfoxide reductases A and B are present in human epidermal melanocytes in the cytosol and in the nucleus

Oxidation of methionine residues by reactive oxygen (ROS) in protein structures leads to the formation of methionine sulfoxide which can consequently lead to a plethora of impaired functionality. The generation of methionine sulfoxide yields ultimately a diastereomeric mixture of the S and R sulfoxides. So far two distinct enzyme families have been identified. MSRA reduces methionine S-sulfoxide, while MSRB reduces the R-diastereomer. It has been shown that these enzymes are involved in regulation of protein function and in elimination of ROS via reversible methionine formation besides protein repair. Importantly, both enzymes require coupling to the NADPH/thioredoxin reductase/thioredoxin electron donor system. In this report, we show for the first time the expression and function of both sulfoxide reductases together with thioredoxin reductase in the cytosol as well as in the nucleus of epidermal melanocytes which are especially sensitive to ROS. Since this cell resides in the basal layer of the epidermis and its numbers and functions are reduced upon ageing and for instance also in depigmentation processes, we believe that this discovery adds an intricate repair mechanism to melanocyte homeostasis and survival.     

Laccase-catalyzed polymerization of tyrosine-containing peptides

Laccase-catalyzed polymerization of tyrosine and tyrosine-containing peptides was studied in the presence and absence of ferulic acid (FA). Advanced spectroscopic methods such as MALDI-TOF MS, EPR, FTIR microscopy and HPLC-fluorescence, as well as more conventional analytical tools: oxygen consumption measurements and SDS/PAGE were used in the reaction mechanism studies. Laccase was found to oxidize tyrosine and tyrosine-containing peptides, with consequent polymerization of the compounds. The covalent linkage connecting the compounds was found to be an ether bond. Only small amounts of dityrosine bonds were detected in the polymers. When FA was added to the reaction mixtures, it was found to be incorporated into the polymer structure. Thus, in addition to homopolymers, different heteropolymers containing two or four FA residues were formed in the reactions.     

Soil CO2 efflux of two silver birch clones exposed to elevated CO2 and O3 levels during three growing seasons

In the present open‐top chamber experiment, two silver birch clones (Betula pendula Roth, clone 4 and clone 80) were exposed to elevated levels of carbon dioxide (CO₂) and ozone (O₃), singly and in combination, and soil CO₂ efflux was measured 14 times during three consecutive growing seasons (1999–2001). In the beginning of the experiment, all experimental trees were 7 years old and during the experiment the trees were growing in sandy field soil and fertilized regularly. In general, elevated O₃ caused soil CO₂ efflux stimulation during most measurement days and this stimulation enhanced towards the end of the experiment. The overall soil respiration response to CO₂ was dependent on the genotype, as the soil CO₂ efflux below clone 80 trees was enhanced and below clone 4 trees was decreased under elevated CO₂ treatments. Like the O₃ impact, this clonal difference in soil respiration response to CO₂ increased as the experiment progressed. Although the O₃ impact did not differ significantly between clones, a significant time × clone × CO₂× O₃ interaction revealed that the O₃‐induced stimulation of soil respiration was counteracted by elevated CO₂ in clone 4 on most measurement days, whereas in clone 80, the effect of elevated CO₂ and O₃ in combination was almost constantly additive during the 3‐year experiment. Altogether, the root or above‐ground biomass results were only partly parallel with the observed soil CO₂ efflux responses. In conclusion, our data show that O₃ impacts may appear first in the below‐ground processes and that relatively long‐term O₃ exposure had a cumulative effect on soil CO₂ efflux. Although the soil respiration response to elevated CO₂ depended on the tree genotype as a result of which the O₃ stress response might vary considerably within a single tree species under elevated CO₂, the present experiment nonetheless indicates that O₃ stress is a significant factor affecting the carbon cycling in northern forest ecosystems.     

Characteristics of the interaction of the ferritin repressor protein with the iron-responsive element

Summary The iron-responsive regulation of ferritin mRNA translation is mediated by the specific interaction of the ferritin repressor protein (FRP) with the iron-responsive element (IRE), a highly conserved 28-nucleotide sequence located in the 5 untranslated region of ferritin mRNAs. The IRE alone is necessary and sufficient to confer repression of translation by FRP upon a heterologous message, chloramphenicol acetyltransferase, in an in vitro translation system. The activity of FRP is sensitive to iron in vivo. Cytoplasmic extracts of rabbit kidney cells show reduction of FRP activity when grown in the presence of iron, as detected by RNA band shift assay. Using a nitrocellulose filter binding assay to examine the interaction of FRP with the IRE in more detail, we find that purified FRP has a single high-affinity binding site for the IRE with aK _d of 20–50 pM. Hemin pretreatment decreases the total amount of FRP which can bind to the IRE. This effect is dependent on hemin concentration. Interestingly, the FRP which remains active at a given hemin concentration binds to the IRE with the same high affinity as untreated FRP. A variety of hemin concentrations were examined for their effect on preformed FRP/IRE complexes. All hemin concentrations tested resulted in rapid complex breakdown. The final amount of complex breakdown corresponds to the concentration of hemin present in the reaction. The effect of hemin on FRP activity suggests that a specific hemin binding site exists on FRP.Abbreviations IRE iron-responsive element - FRP ferritin repressor protein - CAT chloramphenicol acetyltransferase - ORF open reading frame     

Emissions of greenhouse gases from ponds constructed for nitrogen removal

Methane and carbon dioxide emission from three constructed ponds were monitored during an annual cycle. Water temperature was a good predictor of methane emission in all three ponds. In the most intensively studied pond, nitrate concentration in the bottom water could further explain the amount of methane emitted. When water temperature exceeded 15 °C between 1 and 54 mg, CH₄ m⁻² h⁻¹ was emitted on all occasions, while at temperatures below 10 °C, less than 0.6 mg CH₄ m⁻² h⁻¹ was emitted. The flux of carbon dioxide differed between the ponds and no consistent patterns were found. In a laboratory study at 20 °C, we showed that high, but naturally occurring, nitrate concentrations (8 and 16 mg NO₃⁻–N l⁻¹) constrained the production of methane compared to the treatment with no nitrate addition. Nitrous oxide production was positively correlated with nitrate concentration. Carbon dioxide production was highest at the highest nitrate concentration, which indicates that increased nitrate loading on ponds and wetlands will stimulate organic matter decomposition rates. Our conclusion is that these ponds constructed for nitrate removal emit greenhouse gases comparable to lakes in the temperate region.