Methylglyoxal (MG) (pyruvaldehyde) is a reactive carbonyl compound produced in glycolysis. MG can form covalent adducts on proteins resulting in advanced glycation end products that may alter protein function. Here we report that MG covalently modifies the mitochondrial permeability transition pore (PTP), a high conductance channel involved in the signal transduction of cell death processes. Incubation of isolated mitochondria with MG for a short period of time (5 min), followed by removal of excess free MG, prevented both ganglioside GD3-and Ca 2؉ -induced PTP opening and the ensuing membrane depolarization, swelling, and cytochrome c release. Under these conditions MG did not significantly interfere with mitochondrial substrate transport, respiration, or oxidative phosphorylation. The suppression of permeability transition was reversible following extended incubation in MG-free medium. Of the 29 physiological carbonyl and dicarbonyl compounds tested only MG and its analogue glyoxal were able to specifically alter the behavior of the PTP. Using a set of arginine-containing peptides, we found that the major MG-derived arginine adduct formed, following a short time exposure to MG, was the 5-hydro-5-methylimidazol-4-one derivative. These findings demonstrate that MG rapidly modifies the PTP covalently and stabilizes the PTP in the closed conformation. This is probably due to the formation of an imidazolone adduct on an arginine residue involved in the control of PTP conformation (Linder, M. D., Morkunaite-Haimi, S., Kinnunen, P. J. K., Bernardi, P., and Eriksson, O. (2002) J. Biol. Chem. 277, 937-942). We deduce that the permeability transition constitutes a potentially important physiological target of MG.

Chemical modification of mitochondria with the arginine-specific reagents phenylglyoxal (PGO) and 2,3-butanedione (BAD) decreases the Ca 2؉ sensitivity of the permeability transition pore (PTP) and stabilizes it in the closed conformation (Eriksson, O., Fontaine, E., and Bernardi, P. (1998) J. Biol. Chem. 273, 12669 -12674). Unexpectedly, modification of mitochondria with the arginine-specific reagent p-hydroxyphenylglyoxal (OH-PGO) resulted instead in PTP opening. Sequential modification with OH-PGO and PGO (or BAD) revealed that the effects on the PTP depended on the order of the additions. PTP opening was observed when OH-PGO preceded, and PTP closing was observed when OH-PGO followed, the addition of PGO (or BAD). The differential effects of OH-PGO and PGO on the PTP open probability (i) were not modified by the conformation-specific ligands of the adenine nucleotide translocase bongkrekate and atractylate; and (ii) were also observed in deenergized mitochondria, indicating that the effect is exerted directly on the PTP. OH-PGO dramatically sensitized PTP opening, which was triggered by depolarization even in the presence of EGTA. These data show that arginine modification modulates the PTP conformation in a ligand-selective fashion and suggest that the effects of OH-PGO, PGO, and BAD are mediated by the same arginine residues. We analyzed the structure of the arginine adducts by matrix-assisted laser desorption ionization and time-of-flight mass spectrometry using a test peptide and N-acetylarginine. The results indicate that both OH-PGO and PGO react with arginine at a stoichiometry of 2:1 and form stable adducts that may be feasible to identify the PTP at the molecular level.The mitochondrial permeability transition is due to the opening of a large channel, the permeability transition pore (PTP 1 ), that permits diffusion of solutes with a molecular mass Ͻ 1500 Da across the inner mitochondrial membrane (1). Opening of the PTP may result in mitochondrial depolarization, swelling, and rupture of the outer membrane causing release of proteins from the intermembrane compartment. These events are under intense investigation in studies of the signal transduction of apoptosis, where the PTP may integrate several stimuli converging on mitochondria (2). Studies on isolated mitochondria have shown that the PTP is controlled by the membrane potential as well as by several ligands. High ⌬⌿, CsA, ADP, H ϩ , and BKA stabilize the closed conformation of the PTP, whereas low ⌬⌿, intramitochondrial Ca 2ϩ , P i , and CATR promote the open conformation (1).The molecular machinery of the PTP is not understood in great detail. It has been proposed that the PTP is composed of the ANT, the outer membrane VDAC, and mitochondrial cyclophilin (CyP-D), which by interactions with proteins of the Bcl-2 family would assemble to a pore-forming complex (3). This model has grown out of the findings that the ANT as well as the Bcl-2 proapoptotic homologue Bax behave as ion channels in model membranes (4,5) and that the ANT inhibitors BKA...

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Phenolic compounds are widespread in berries and determine their antimicrobial activity. The aim of our study was to establish the amounts of phenolic compounds and the anthocyanin composition in berries of four Ribes species, and to evaluate the effect of berry extracts on the growth of common Gram-positive and Gram-negative bacteria, and also yeasts isolated from food processing plants. The phenolic content and anthocyanin composition were estimated spectrometrically and by HPLC, respectively. The highest amount of phenolic compounds, and also anthocyanins, was found in extracts of R. aureum ‘Corona’. The anthocyanin content was the lowest in berries of R. aureum Au Gs-5, with equal amounts of delphinidins and cyanidins. Delphinidins were predominant (68.6%) in berries of R. nigrum ‘Ben Tirran’, while cyanidins dominated in R. uva-crispa. The berry extracts of R. aureum Au Gs-5 and R. uva-crispa ‘Lûðiai’ had the largest growth-suppressing effect on yeasts and most of the bacteria tested. All of the berry extracts suppressed the growth of pathogenic and conditionally pathogenic bacteria. The industrially important Lactococcus lactis was the most resistant to the Ribes berry extracts. There was no correlation between the amount of anthocyanins in the extracts and their antimicrobial properties. Extracts with a lower anthocyanin–to-phenolics ratio more effectively inhibited the growth of bacteria.