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D E Griffiths

Publications and source records attributed to D E Griffiths.

At least 19 recordsLinked to original sources
PubMed1992

Organotin-flavone complexes: a new class of fluorescent probes for F1F0ATPase.

Fluorescent 5-coordinate organotin-flavone complexes of 3-hydroxy-flavone (Hof) and 3,5,7,2',4',-pentahydroxyflavone (morin) are good inhibitors of mitochondrial F1F0ATPase but do not inhibit F1-ATPase and they have been examined as possible fluorescent probes of F1F0ATPase. R2SnX (morin) complexes exhibit low fluorescence enhancement on binding to mitochondrial membranes with no displacement by equimolar tributyltin. In contrast R2SnX (of) complexes exhibit high fluorescence enhancement whose extent is variable and is displacable by equimolar tributyltin. Fluorescence enhancement by R2SnX (of) complexes correlates with the ATPase I50 values. Dialkyltin-3-hydroxy flavone, R2SnX(of), complexes act as a new class of fluorescent probes which titrate the F0 segment of F1F0ATPase.

Animals↗
PubMed1991

Characterization of ATP12, a yeast nuclear gene required for the assembly of the mitochondrial F1-ATPase.

Mitochondrial F1-ATPase is an oligomeric enzyme composed of five distinct subunit polypeptides. The alpha and beta subunits make up the bulk of protein mass of F1. In Saccharomyces cerevisiae both subunits are synthesized as precursors with amino-terminal targeting signals that are removed upon translocation of the proteins to the matrix compartment. Recently, two different complementation groups (G13, G57), consisting of yeast nuclear mutants with defective F1, have been described. Biochemical analyses indicate that the mutational block in both groups of mutants affects a critical step needed for the assembly of the alpha and beta subunits into the F1 oligomer after their transport into mitochondria. In this study the ATP12 gene representative of the nuclear respiratory-deficient mutant of S. cerevisiae (pet) complementation group G57 has been cloned and the encoded product partially characterized. The ATP12 reading frame is 975 base pairs long and codes for a protein of Mr = 36,587. The ATP12 protein is not homologous to the subunits of F1 whose sequences are known, nor does it exhibit significant primary structure similarity to any known protein. In vitro import assays indicate that ATP12 protein is synthesized as a precursor approximately 3 kDa larger than the mature protein. The mitochondrial localization of the protein has been confirmed by Western blot analysis of mitochondrial proteins with an antibody against a hybrid protein expressed from a trpE-ATP12 fusion. Fractionation of mitochondria indicates further that the ATP12 protein is either a minor component of the matrix compartment or is weakly bound to the matrix side of the inner membrane. The molecular weight of the native protein, estimated from its sedimentation properties in sucrose gradients, is at least two times larger than the monomer. This suggests that the ATP12 protein is probably part of a larger complex.

Amino Acid Sequence↗
PubMed1991

Changes in amplitude and latency of the P300 component of the auditory evoked potential with sedative and anaesthetic concentrations of nitrous oxide.

The P300 component of the auditory evoked response was recorded from six subjects whilst they listened via headphones to a series of clicks which were interrupted unpredictably by a tone burst. They were instructed to press a button as quickly as possible after hearing the tone whilst breathing first air and then a series of increasing concentrations of nitrous oxide. Both the amplitude and the latency of the P300 changed in a dose-dependent manner with nitrous oxide, as did minimum reaction time. At nitrous oxide concentrations which prevented recall of any events that occurred whilst breathing the gas, four subjects continued to respond to the tone by pressing the button. In three subjects, the P300 wave was still detectable with a nitrous oxide concentration at which the task was no longer performed. These results show that there is retention of the ability to perform a reaction time task when there is a complete loss of recall of the task. There may be some recognition of an auditory stimulus, as manifest by a P300 wave, albeit reduced greatly in amplitude, in the absence of a motor response to it. The P300, therefore, merits investigation as a tool for studying conscious awareness under anaesthesia.

Anesthesia Recovery Period↗
PubMed1988

DNA sequence analysis of the Olir2-76 and Ossr1-92 alleles of the Oli-2 region of the yeast Saccharomyces cerevisiae. Analysis of related amino-acid substitutions and protein-antibiotic interaction.

Petite deletion mapping helped to generate a fine-structure genetic map of the Oli-2 region of the mitochondrial genome of Saccharomyces cerevisiae. Here we report the DNA sequence analysis of the Oli-2 region from two drug-resistant alleles (Olir2-76 and Ossr1-92) which are located in the gene for subunit-6 of mitochondrial ATPase, in agreement with their genetic locations on the mitochondrial genome. An analysis of the corresponding amino-acid substitutions is also presented in the context of protein-antibiotic interactions.

Adenosine Triphosphatases↗
PubMed1984

Differential inhibition of F0F1-ATPase-catalysed reactions in bovine-heart submitochondrial particles by organotin compounds.

Preincubation of coupled submitochondrial particles with low concentrations of triorganotin compounds results in complete inhibition of the oligomycin-sensitive ATPase activity without any significant effect on the rate of succinate-driven ATP synthesis. The residual ATP synthetic activity is inhibited by oligomycin and uncouplers. The differential inhibition of ATP synthesis and hydrolysis by the triorganotin compounds examined suggests that the two processes are not 'mirror images' of each other, but that they occur through different routes and that the F1F0-ATPase is at least bifunctional.

Adenosine Triphosphatases↗
PubMed1977

Dibutylchloromethyltin chloride, a covalent inhibitor of the adenosine triphosphate synthase complex.

1. The synthesis of dibutylchloromethyltin chloride, a new covalent inhibitor of the mitochondrial ATP synthase [oligomycin-sensitive ATPase (adenosine triphosphatase)] complex is described, together with a method for preparing dibutylchloro[(3)H]methyltin chloride. 2. Studies with the yeast mitochondrial oligomycin-sensitive ATPase complex show that dibutylchloromethyltin chloride inhibits both the membrane-bound enzyme and also the purified Triton X-100-dispersed preparation. 3. F(1)-ATPase is not inhibited even at 500nmol of dibutylchloromethyltin chloride/mg of protein, and the general inhibitory properties are similar to those of triethyltin, oligomycin and dicyclohexylcarbodi-imide, known energy-transfer inhibitors of oxidative phosphorylation. 4. Binding studies with yeast submitochondrial particles show that dibutylchloromethyltin chloride antagonizes the binding of triethyl[(113)Sn]tin, indicating that there is an interaction between the two inhibitor-binding sites. 5. Unlike triethyltin, inhibition by dibutylchloromethyltin chloride is due to a covalent interaction which titrates a component of the inner mitochondrial membrane present at a concentration of 8-9nmol/mg of protein. 6. All of the labelled component can be extracted with chloroform/methanol (2:1, v/v), and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the chloroform/methanol extract indicates that the labelled component has an apparent mol.wt. of 6000-8000. However, t.l.c. reveals the presence of only one labelled component which is lipophilic and non-protein and is distinct from the free inhibitor, mitochondrial phospholipids and the dicyclohexylcarbodi-imide-binding protein (subunit 9). 7. Inhibition of mitochondrial ATPase and oxidative phosphorylation is correlated with specific interaction with a non-protein lipophilic component of the mitochondrial inner membrane which is proposed to be a co-factor or intermediate of oxidative phosphorylation.

Adenosine Triphosphatases↗
PubMed1977

Studies of energy-linked reactions. Inhibition of oxidative phosphorylation by DL-8-methyldihydrolipoate.

1. DL-8-Methyldihydrolipoate was shown to be a potent inhibitor of mitochondrial oxidative phosphorylation and ATP-driven energy-linked reactions. 2. ADP-stimulated respiration utilizing pyruvate + malate and succinate in both ox heart and rat liver mitochondria is inhibited; oxidative phosphorylation using pyruvate + malate, succinate and ascorbate + NNN'N'-tetramethyl-p-phenylenediamine as substrates is also inhibited; uncoupler-stimulated respiration is unaffected regardless of the substrate used. 3. Mitochondrial oligomycin-sensitive adenosine triphosphatase is inhibited in both the membrane-bound form and the purified detergent-dispersed preparation. 4. ATP-driven transhydrogenase and the ATP-driven energy-linked reduction of NAD+ by succinate in ox heart submitochondrial particles are inhibited, whereas the respiratory-chain-driven transhydrogenase is unaffected. 5. DL-8-Methyl-lipoate has no immediate effect on the above reactions, demonstrating the requirement for the reduced form for inhibition. 6. The inhibitory properties of DL-8-methyldihydrolipoate are analogous to those of oligomycin and provide further evidence of a role for lipoic acid in oxidative phosphorylation.

Adenosine Triphosphatases↗
PubMed1977

The isolation and genetic characterization of extrachromosomal chloramphenicol and oligomycin-resistant mutants from the petite-negative yeast Kluyveromyces lactis.

Spontaneous mutants of the petite-negative yeast Kluyveromyces lactis, resistant to the antibiotics chloramphenicol and oligomycin, were isolated and genetically characterized. Three chloramphenicol-resistant mutants showed non-Mendelian inheritance when crossed to sensitive parents. Of 5 oligomycin-resistant strains studied, three exhibited resistance due to the presence of an extrachromosomal mutation. The resistance of the other two deriving from a nuclear and recessive mutation. When two factor crosses in trans configuration were performed between two of the chloramphenicol and the five oligomycin-resistant mutants a polarity in recombination was observed with a predominance of sensitive (OSCS) over resistant (ORCR) reciprocal recombinants. Allelism tests carried out among the oligomycin-resistant mutants indicated the presence of at least two distinct extrachromosomal regions responsible for the resistance.

Chloramphenicol↗
PubMed1977

Studies of energy-linked reactions. Localization of the site of action of trialkyltin in yeast mitochondria.

Ligand-binding studies with labelled triethyltin on yeast mitochondrial membranes showed the presence of high-affinity sites (KD = 0.6 micronM; 1.2 +/- 0.3 nmol/mg of protein) and low-affinity sites (KD less than 45 micronM; 70 +/- 20 nmol/mg of protein). The dissociation constant of the high-affinity site is in good agreement with the concentration of triethyltin required for inhibition of mitochondrial ATPase (adenosine triphosphatase) and oxidative phosphorylation. The high-affinity site is not competed for by oligomycin or venturicidin, indicating that triethyltin reacts at a different site from these inhibitors of oxidative phosphorylation. Fractionation of the mitochondrial membrane shows a specific association of the high-affinity sites with the ATP synthase complex. During purification of ATP synthase (oligomycin-sensitive ATPase) there is a 5-6-fold purification of oligomycin- and triethyltin-sensitive ATPase activity concomitant with a 7-9-fold increase in high-affinity triethyltin-binding sites. The purified yeast oligomycin-sensitive ATPase complex contains approximately six binding sites for triethyltin/mol of enzyme complex. It is concluded that specific triethyltin-binding sites are components of the ATP synthase complex, which accounts for the specific inhibition of ATPase and oxidative phosphorylation by triethyltin.

Adenosine Triphosphatases↗