Coupling mechanisms in anionic substrate transport across the inner membrane of rat-liver mitochondria

The translocation of P_i, malate, -oxoglutarate, and citrate across the inner membrane of rat-liver mitochondria has been studied. Investigation on the effect of pH on anionic substrate translocation across the mitochondrial membrane shows that their distribution across the inner membrane can be governed by transmembrane pH difference. However, evidence is presented that the translocation of P_i, but not that of malate, -oxoglutarate, or citrate can bedirectly coupled to an OH^– counterflux (H₂PO ₄ ^– –OH^– exchange-diffusion). and malate-tricarboxylate exchange-diffusion reactions is directly demonstrated. The study of the effect of uncouplers on the efflux from mitochondria of substrate anions, in the absence of counteranion, and on the anion exchange-diffusions shows that uncouplers act in at least two ways: they promote the efflux of P_i from mitochondria and inhibitdirectly the exchange-diffusion reactions. The kinetics of this inhibition are described. These results are discussed in the light of previous work on the effect of uncouplers on the distribution of substrate anions across the inner membrane of isolated mitochondria. Coupling mechanisms in substrate anion translocation and aspects of the energetics of anion translocation are discussed.     

Topological and functional characterization of the F0I subunit of the membrane moiety of the mitochondrial H+-ATP synthase

Using isolated polypeptides of the F0 sector of bovine heart mitochondrial H+-ATPase, antisera were developed detecting specifically two components of F0. These two components were identified as F0I and oligomycin-sensitivity-conferring protein (OSCP) respectively. Both F0I and OSCP were digested by mild trypsin treatment of submitochondrial particles depleted of the catalytic part of H+-ATPase (USMP). Proteolysis was largely prevented by binding of F1 to F0. Proteolysis of F0I resulted in the formation of three immunoreactive, membrane-bound fragments of apparently 26 kDa, 25.5 kDa and 18 kDa, respectively, indicating that F0I contains trypsin-accessible Arg or Lys residues located close to the end and the middle part of the protein, respectively, which are in intimate contact with F1. Digestion of USMP with trypsin resulted in depression of passive H+ conduction through F0 which could be ascribed to proteolysis of F0I.     

Bacterial biofilm formation inhibitory activity revealed for plant derived natural compounds

Use of herbal plant remedies to treat infectious diseases is a common practice in many countries in traditional and alternative medicine. However to date there are only few antimicrobial agents derived from botanics. Based on microbiological screening tests of crude plant extracts we identified four compounds derived from Krameria, Aesculus hippocastanum and Chelidonium majus that showed a potentially interesting antimicrobial activity. In this work we present an in depth characterization of the inhibition activity of these pure compounds on the formation of biofilm of Staphylococcus aureus as well as of Staphylococcus epidermidis strains. We show that two of these compounds possess interesting potential to become active principles of new drugs.     

Dysfunctions of cellular oxidative metabolism in patients with mutations in the NDUFS1 and NDUFS4 genes of complex I

The pathogenic mechanism of a G44A nonsense mutation in the NDUFS4 gene and a C1564A mutation in the NDUFS1 gene of respiratory chain complex I was investigated in fibroblasts from human patients. As previously observed the NDUFS4 mutation prevented complete assembly of the complex and caused full suppression of the activity. The mutation (Q522K replacement) in NDUFS1 gene, coding for the 75-kDa Fe-S subunit of the complex, was associated with (a) reduced level of the mature complex, (b) marked, albeit not complete, inhibition of the activity, (c) accumulation of H(2)O(2) and O(2)(.-) in mitochondria, (d) decreased cellular content of glutathione, (e) enhanced expression and activity of glutathione peroxidase, and (f) decrease of the mitochondrial potential and enhanced mitochondrial susceptibility to reactive oxygen species (ROS) damage. No ROS increase was observed in the NDUFS4 mutation. Exposure of the NDUFS1 mutant fibroblasts to dibutyryl-cAMP stimulated the residual NADH-ubiquinone oxidoreductase activity, induced disappearance of ROS, and restored the mitochondrial potential. These are relevant observations for a possible therapeutical strategy in NDUFS1 mutant patients.