Redox-linked proton transfer by cytochrome c oxidase

The respiratory chain is found in the inner mitochondrial membrane of higher organisms and in the plasma membrane of many bacteria. It consists of several membrane-spanning enzymes, which conserve the energy that is liberated from the degradation of food molecules as an electrochemical proton gradient across the membrane. The proton gradient can later be utilized by the cell for different energy requiring processes, e.g. ATP production, cellular motion or active transport of ions. The difference in proton concentration between the two sides of the membrane is a result of the translocation of protons by the enzymes of the respiratory chain, from the negatively charged (N-side) to the positively charged side (P-side) of the lipid bilayer, against the proton concentration gradient. The endergonic proton transfer is driven by the flow of electrons through the enzymes of the respiratory chain, from low redox-potential electron donors to acceptors of higher potential, and ultimately to oxygen. Cytochrome c oxidase is the last enzyme in the respiratory chain and catalyzes the reduction of dioxygen to water. The redox reaction is coupled to proton transport across the membrane by a yet unresolved mechanism. Cytochrome c oxidase has two proton-conducting pathways through which protons are taken up to the interior part of the enzyme from the N-side of the membrane. The K-pathway transfers merely substrate protons, which are consumed in the process of water formation at the catalytic site. The D-pathway transfers both substrate protons and protons that are pumped to the P-side of the membrane. This thesis focuses on the role of two conserved amino acids in proton translocation by cytochrome c oxidase, glutamate 278 and tryptophan 164. Glu278 is located at the end of the D-pathway and is thought to constitute the branching point for substrate and pumped protons. In this work, it was shown that although Glu278 has an important role in the proton transfer mechanism, its presence is not an obligatory requirement. Alternative structural solutions in the area around Glu278, much like the ones present in some distantly related heme-copper oxidases, could in the absence of Glu278 support the formation of a long hydrogen-bonded water chain through which proton transfer from the D-pathway to the catalytic site is possible. The other studied amino acid, Trp164, is hydrogen bonded to the ∆-propionate of heme a3 of the catalytic site. Mutation of this amino acid showed that it may be involved in regulation of proton access to a proton acceptor, a pump site, from which the proton later is expelled to the P-side of the membrane. The ion pair that is formed by the ∆-propionate of heme a3 and arginine 473 is likely to form a gate-like structure, which regulates proton mobility to the P-side of the membrane. The same gate may also be part of an exit path through which water molecules produced at the catalytically active site are removed towards the external side of the membrane. Time-resolved optical and electrometrical experiments with the Trp164 to phenylalanine mutant revealed a so far undetected step in the proton pumping mechanism. During the A to PR transition of the catalytic cycle, a proton is transferred from Glu278 to the pump site, located somewhere in the vicinity of the ∆-propionate of heme a3. A mechanism for proton pumping by cytochrome c oxidase is proposed on the basis of the presented results and the mechanism is discussed in relation to some relevant experimental data. A common proton pumping mechanism for all members of the heme-copper oxidase family is moreover considered.

Utveckling av en ny modul i WM-datas arbetsorderhanteringssystem för registrering av snöröjning respektive sandning

WM-data tillhandahåller en lösning för mobil arbetsorderhantering. Lösningen innehåller planeringsverktyg med ärenden, personal och karta och möjlighet att leverera arbetsorder till fältpersonal. Den innehåller dessutom handhållna mobilenheter, vilka fältpersonalen använder för att ta emot aktuella arbetsorder.Vissa kunder önskar ytterligare en modul i denna mobila lösning. Den nya modulen skall användas för att kontinuerligt rapportera in genomförda åtgärder i snöröjningssammanhang.Detta examensarbete går ut på att utveckla den här modulen för att registrera snöröjning respektive sandning. Applikationen exekveras på en PDA, en s�� kallad Pocket PC med Windows Mobile 5.0. Anledningen till att just Windows Mobile 5.0 skall användas är att detta operativsystem har inbyggt stöd för GPS-positionering. Ett önskemål av WM-data var att detta inbyggda stöd skulle unders��kas, rapporteras och användas i applikationen.Resultatet blev en applikation där man via en Pocket PC kontinuerligt sparar undan fordons GPS-position, arbets��tgärd samt tidpunkterna när GPS-positionerna togs. Denna information är sedan anpassad för att kunna skickas vidare via GPRS till en befintlig databasserver under vissa tidsintervaller.Applikationen utvecklades med hjälp av programmeringsspråket C Sharp (C#) i Visual Studio 2005.

The value of military oath in civil wars in Rome: The example of conflict 49-45 BC

During the civil war between Caesar and Pompey, the military oath which binds the soldier to his army is often openly violated. Yet despite this offense, commanders of armed struggle require recursively the oath to their men. Admittedly, this ritual act seems ineffective given the many desertions and mutinies identified, but military leaders use its symbolic and sacred meaning to legitimize one hand their “anti-republican” actions, on the other armies fighting in a context deemed impius.

Le “brigand lusitanien” reconsidéré. Analyse du problème de la terre chez Appien

L’étude du “brigandage lusitanien” a donné lieu à une importante activité de recherche depuis la fin du XIXe siècle. Pour autant, et malgré une inflexion progressive de l’historiographie moderne vers une approche plus nuancée de l’origine du phénomène, le problème de la terre reste encore aujourd’hui au centre des préoccupations de nombre d’historiens et archéologiques. À partir d’une discussion serrée des principaux passages de Tite-Live, Diodore et surtout Appien, relatifs à la relation que d’aucuns ont voulu établir entre manque et/ou pauvreté de la terre et développement du brigandage chez les Lusitaniens, il est propos�� une critique de l’interprétation socio-économique.

Adult-type hypolactasia: Genotype-phenotype correlation

Adult-type hypolactasia (primary lactose malabsorption, lactase non-persistence) is the most common enzyme deficiency worldwide, and manifests with symptoms of lactose intolerance such as abdominal pain, gas formation and diarrhea. In humans with adult-type hypolactasia, lactase activity is high at birth, but declines during childhood to about one-tenth of the activity at birth. In 2002, a one base polymorphism C/T-13910, located 14 kilobases from the starting codon of the lactase-phlorizin hydrolase (LPH) gene was observed to be associated with the persistence of lactase activity. The T-13910 allele (C/T-13910 and T/T-13910 genotypes) associates with persistence of lactase activity throughout life, whereas the C/C-13910 genotype associates with adult-type hypolactasia. In this thesis work, the timing and mechanism of decline of lactase enzyme activity during development was studied using the C/T-13910 polymorphism as a molecular marker. We observed an excellent correlation between low lactase activity and the C/C-13910 genotype in all subjects > 12 years of age, irrespective their ethnicity. In children of African origin, the lactase activity declined somewhat earlier than among Finnish children. Furthermore, we observed an increasing imbalance in the relative lactase mRNA expression from the C-13910 and T-13910 alleles in Finnish children beginning from five years of age. The genetic test for adult-type hypolactasia showed a sensitivity of 93% and a specificity of 100% in the Finnish children and adolescents > 12 years of age. The relation of milk consumption and the milk-related abdominal complaints to the C/T-13910 genotypes associated with lactase persistence/non-persistence was studied by a questionnaire-based approach in > 2100 Finns. Both Finnish children and adults with the C/C-13910 genotype consumed significantly less dairy products compared to those with the C/T-13910 and T/T-13910 genotypes. Flatulence was the only of the abdominal symptoms of lactose intolerance that subjects with the C/C-13910 genotype reported significantly more often than those with the C/T-13910 and T/T-13910 genotypes. A minor proportion (<10%) of subjects with the C/C-13910 genotype, nevertheless, reported drinking milk without any symptoms afterwards. There was no association between cow's milk allergy starting as a newborn and adult-type hypolactasia. In an association study an increased risk of colorectal cancer was observed among those with molecular diagnosis of adult-type hypolactasia. It warrants further studies to clarify whether the increased risk observed in the Finnish population is associated with lactose or decreased intake of dairy products in these subjects.