Amyloid Formation by the Pro-Inflammatory S100A8/A9 Proteins in the Ageing Prostate

BACKGROUND: The conversion of soluble peptides and proteins into polymeric amyloid structures is a hallmark of many age-related degenerative disorders, including Alzheimer's disease, type II diabetes and a variety of systemic amyloidoses. We report here that amyloid formation is linked to another major age-related phenomenon--prostate tissue remodelling in middle-aged and elderly men. METHODOLOGY/PRINCIPAL FINDINGS: By using multidisciplinary analysis of corpora amylacea inclusions in prostate glands of patients diagnosed with prostate cancer we have revealed that their major components are the amyloid forms of S100A8 and S100A9 proteins associated with numerous inflammatory conditions and types of cancer. In prostate protease rich environment the amyloids are stabilized by dystrophic calcification and lateral thickening. We have demonstrated that material closely resembling CA can be produced from S100A8/A9 in vitro under native and acidic conditions and shows the characters of amyloids. This process is facilitated by calcium or zinc, both of which are abundant in ex vivo inclusions. These observations were supported by computational analysis of the S100A8/A9 calcium-dependent aggregation propensity profiles. We found DNA and proteins from Escherichia coli in CA bodies, suggesting that their formation is likely to be associated with bacterial infection. CA inclusions were also accompanied by the activation of macrophages and by an increase in the concentration of S100A8/A9 in the surrounding tissues, indicating inflammatory reactions. CONCLUSIONS/SIGNIFICANCE: These findings, taken together, suggest a link between bacterial infection, inflammation and amyloid deposition of pro-inflammatory proteins S100A8/A9 in the prostate gland, such that a self-perpetuating cycle can be triggered and may increase the risk of malignancy in the ageing prostate. The results provide strong support for the prediction that the generic ability of polypeptide chains to convert into amyloids could lead to their involvement in an increasing number of otherwise apparently unrelated diseases, particularly those associated with ageing.Original Publication:Kiran Yanamandra, Oleg Alexeyev, Vladimir Zamotin, Vaibhav Srivastava, Andrei Shchukarev, Ann-Christin Brorsson, Gian Gaetano Tartaglia, Thomas Vogl, Rakez Kayed, Gunnar Wingsle, Jan Olsson, Christopher M Dobson, Anders Bergh, Fredrik Elgh and Ludmilla A Morozova-Roche, Amyloid formation by the pro-inflammatory S100A8/A9 proteins in the ageing prostate., 2009, PloS one, (4), 5, e5562.http://dx.doi.org/10.1371/journal.pone.000556

Routine sample preparation and HPLC analysis for ascorbic acid (vitamin C) determination in wheat plants and Arabidopsis leaf tissues

Plants have developed various mechanisms to protect themselves against oxidative stress. One of the most important non-enzymatic antioxidants is ascorbic acid. There is thus a need for a rapid, sensitive method for the analysis of the reduced and oxidised forms of ascorbic acid in crop plants. In this paper a simple, economic, selective, precise and stable HPLC method is presented for the detection of ascorbate in plant tissue. The sensitivity, the short retention time and the simple isocratic elution mean that the method is suitable for the routine quantification of ascorbate in a high daily sample number. The method has been found to be better than previously reported methods, because of the use of an economical, readily available mobile phase, UV detection and the lack of complicated extraction procedures. The method has been tested on Arabidopsis plants with different ascorbate levels and on wheat plants during Cd stress

S-ADENOSYL-L-METHIONINE-THEOBROMINE 1-N-METHYLTRANSFERASE, AN ENZYME CATALYZING THE SYNTHESIS OF CAFFEINE IN COFFEE

This paper presents data on the purification and N-terminal sequence of S-adenosyl-L-methionine: theobromine 1-N-methyltransferase (STM), the enzyme responsible for the methylation of theobromine leading to caffeine formation in coffee. STM was purified from developing endosperms of immature fruits by DEAF-cellulose, hydrophobic interaction and affinity chromatography, using S-adenosyl-L-homocysteine as a ligand. The enzyme showed an apparent M(r) of ca 54 000 and ca 60 000 determined by gel filtration and SDS-PAGE, respectively. A pI of 5.1 was obtained by liquid chromatofocusing and 4.8 by isoelectrofocusing in polyacrylamide gel electrophoresis. Using theobromine as substrate, the K-m value for S-adenosyl-L-methionine was 10 mu M, being competitively inhibited by S-adenosyl-L-homocysteine (K-i = 4.6 mu M). STM is a bifunctional enzyme since it also methylated 7-methylxanthine, the immediate precursor of theobromine in the caffeine biosynthesis pathway. The specific activity of STM with 7-methylxanthine was ca 55% of that determined with theobromine. The K-m values obtained for theobromine and 7-methylxanthine were 0.196 and 0.496 mM, respectively. STM was also purified from leaves using the same procedures used for endosperms, plus an additional chromatography on a Mono Q column; theobromine was used as substrate. Finally, the N-terminal sequence for the first 20 amino acids was obtained for STM purified from endosperms. No similarities were found with other methyltransferase sequences or other known proteins.3761577158

Cold acclimation and photoinhibition of photosynthesis in Scots pine

Cold acclimation of Scots pine did not affect the susceptibility of photosynthesis to photoinhibition. Cold acclimation did however cause a suppression of the rate of CO2 uptake, and at given light and temperature conditions a larger fraction of the photosystem II reaction centres were closed in cold-acclimated than in nonacclimated pine. Therefore, when assayed at the level of photosystem II reaction centres, i.e. in relation to the degree of photosystem closure, cold acclimation caused a significant increase in resistance to photoinhibition; at given levels of photosystem II closure the resistance to photoinhibition was higher after cold acclimation. This was particularly evident in measurements at 20°C. The amounts and activities of the majority of analyzed active oxygen scavengers were higher after cold acclimation. We suggest that this increase in protective enzymes and compounds, particularly superoxide dismutase, ascorbate peroxidase, glutathione reductase and ascorbate of the chloroplasts, enables Scots pine to avoid excessive photoinhibition of photosynthesis despite partial suppression of photosynthesis upon cold acclimation. An increased capacity for light-induced de-epoxidation of violaxanthin to zeaxanthin upon cold acclimation may also be of significance