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Cytochrome c oxidase is a membrane protein of the respiratory chain that consumes protons and molecular oxygen to produce water and uses the resulting energy to pump protons across the membrane. Our molecular dynamics simulations with an excess proton located at different positions in one of the proton-conducting channels, the K-channel, show a clear dependence of the number of water molecules inside the channel on the proton position. A higher hydration level facilitates the formation of hydrogen-bonded chains along which proton transfer can occur. However, a sufficiently high hydration level for such proton transport is observed only when the excess proton is located above S365, i.e., the lower third of the channel. From the channel entrance up to this point, proton transport is via water molecules as proton carriers. These hydronium ions move with their surrounding water molecules, up to K362, filling and widening the channel. The conformation of K362 depends on its own protonation state and on the hydration level, suggesting its role to be proton transport from a hydronium ion at the height of K362 to the upper part of the channel via a conformational change. The protonation-dependent conformational dynamics of E101 at the bottom of the channel renders proton transfer via E101 unlikely. Instead, its role is rather that of an amplifier of H96’s proton affinity, suggesting H96 as the initial proton acceptor.

Thymine DNA Glycosylase (TDG) is an enzyme of the base excision repair mechanism and removes damaged or mispaired bases from DNA via hydrolysis of the glycosidic bond. Specificity is of high importance for such a glycosylase, so as to avoid the damage of intact DNA. Among the substrates reported for TDG are mispaired uracil and thymine but also formyl-cytosine and carboxyl-cytosine. Methyl-cytosine and hydroxylmethyl-cytosine are, in contrast, not processed by the TDG enzyme. We have in this work employed molecular dynamics simulations to explore the conformational dynamics of DNA carrying a formyl-cytosine or carboxyl-cytosine and compared those to DNA with the non-cognate bases methyl-cytosine and hydroxylmethyl-cytosine, as amino and imino tautomers. Whereas for the mispairs a wobble conformation is likely decisive for recognition, all amino tautomers of formyl-cytosine and carboxyl-cytosine exhibit the same Watson���Crick conformation, but all imino tautomers indeed form wobble pairs. The conformational dynamics of the amino tautomers in free DNA do not exhibit differences that could be exploited for recognition, and also complexation to the TDG enzyme does not induce any alteration that would indicate preferable binding to one or the other oxidised methyl-cytosine. The imino tautomers, in contrast, undergo a shift in the equilibrium between a closed and a more open, partially flipped state, towards the more open form upon complexation to the TDG enzyme. This stabilisation of the more open conformation is most pronounced for the non-cognate bases methyl-cytosine and hydroxyl-cytosine and is thus not a likely mode for recognition. Moreover, calculated binding affinities for the different forms indicate the imino forms to be less likely in the complexed DNA. These findings, together with the low probability of imino tautomers in free DNA and the indifference of the complexed amino tautomers, suggest that discrimination of the oxidised methyl-cytosines does not take place in the initial complex formation.

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Proton transfer reactions are one of the most fundamental processes in biochemistry. We present a simplistic approach for estimating proton transfer probabilities in a membrane protein, cytochrome c oxidase. We combine short molecular dynamics simulations at discrete protonation states with a Monte Carlo approach to exchange between those states. Requesting for a proton transfer the existence of a hydrogen-bonded connection between the two source and target residues of the exchange, restricts the acceptance of transfers to only those in which a proton-relay is possible. Together with an analysis of the hydrogen-bonded connectivity in one of the proton-conducting channels of cytochrome c oxidase, this approach gives insight into the protonation dynamics of the hydrogen-bonded networks. The connectivity and directionality of the networks are coupled to the conformation of an important protein residue in the channel, K362, rendering proton transfer in the entire channel feasible in only one of the two major conformations. Proton transport in the channel can thus be regulated by K362 not only through its possible role as a proton carrier itself, but also by allowing or preventing proton transport via water residues.

Experimental and theoretical methods were used to study two mixtures, 1-hexanol with n-hexane and cis-3-hexen-1-ol with n-hexane. The experimental technics were used to determine excess molar volume and viscosity deviations in the temperature range from 288.15K to 318.15K. The data of excess molar volume indicate similar behavior of the two alcohols, 1-hexanol and cis-3-hexen-1-ol in mixtures with n-hexane. The experimental data are in accordance with molecular dynamics simulation of these mixtures, showing similar number of hydrogen bonds in the two mixtures. The experimental data of viscosity deviations show difference of the two mixtures, where the deviation is significantly larger for cis-3-hexen-1-ol and n-hexane mixture, indicating stronger interactions within cis-3-hexen-1-ol and n-hexane binary mixture. These experimental data are in accordance with quantum chemical calculations on interactions energies. Namely, calculated interaction energies show that interaction between single and double bonds are stronger, than interactions between two single bonds. Eksperimentalne i teorijske metode korišćene su za proučavanje dve smeše, 1-heksanola sa n-heksanom i cis-3-heksen-1-ola sa n-heksanom. Eksperimentalne tehnike su korišćene za određivanje dopunskih zapremina i promene viskoznosti za temperaturni opseg od 288.15K do 318.15K. Podaci o dopunskoj molarnoj zapremini ukazuje na slično ponašanje dva alkohola, 1-heksanola i cis-3-heksen-1-ola u smešama sa n-heksanom. Eksperimentalni podaci su u skladu sa simulacijom molekularne dinamike ovih smeša, pokazujući sličan broj vodoničnih veza u dve smeše. Eksperimentalni podaci o promeni viskoznosti pokazuju značajnu razliku između dve smeše, odstupanje je značajno veće za smešu cis-3-heksen-1-ol i n-heksan, što ukazuje na jaču interakciju u binarnoj smeši cis-3-heksen-1-ola i nheksana. Ovi eksperimentalni podaci su u skladu sa kvantno-hemijskim proračunima energija interakcija na molekulskom nivou. Naime, izračunate energije interakcije pokazuju da je interakcija između jednostrukih i dvostrukih veza jača, nego interakcija između dve jednostruke veze. Related to: [http://cer.ihtm.bg.ac.rs/handle/123456789/3291]

Experimental and theoretical methods were used to study two mixtures, 1-hexanol with nhexane and cis-3-hexen-1-ol with n-hexane. The experimental technics were used to determine excess molar volume and viscosity deviations in the temperature range from 288.15K to 318.15K. The data of excess molar volume indicate similar behavior of the two alcohols, 1-hexanol and cis-3-hexen-1-ol in mixtures with n-hexane. The experimental data are in accordance with molecular dynamics simulation of these mixtures, showing similar number of hydrogen bonds in the two mixtures. The experimental data of viscosity deviations show difference of the two mixtures, where the deviation is significantly larger for cis-3- hexen-1-ol and n-hexane mixture, indicating stronger interactions within cis-3-hexen-1-ol and n-hexane binary mixture. These experimental data are in accordance with quantum chemical calculations on interactions energies. Namely, calculated interaction energies show that interaction between single and double bonds are stronger, than interactions between two single bonds. Eksperimentalne i teorijske metode korišćene su za proučavanje dve smeše, 1-heksanola sa n-heksanom i cis-3-heksen-1-ola sa n-heksanom. Eksperimentalne tehnike su korišćene za određivanje dopunskih zapremina i promene viskoznosti za temperaturni opseg od 288.15K do 318.15K. Podaci o dopunskoj molarnoj zapremini ukazuje na slično ponašanje dva alkohola, 1-heksanola i cis-3-heksen-1-ola u smešama sa n-heksanom. Eksperimentalni podaci su u skladu sa simulacijom molekularne dinamike ovih smeša, pokazujući sličan broj vodoničnih veza u dve smeše. Eksperimentalni podaci o promeni viskoznosti pokazuju značajnu razliku između dve smeše, odstupanje je značajno veće za smešu cis-3-heksen-1- ol i n-heksan, što ukazuje na jaču interakciju u binarnoj smeši cis-3-heksen-1-ola i nheksana. Ovi eksperimentalni podaci su u skladu sa kvantno-hemijskim proračunima energija interakcija na molekulskom nivou. Naime, izračunate energije interakcije pokazuju da je interakcija između jednostrukih i dvostrukih veza jača, nego interakcija između dve jednostruke veze. Related to: [http://cer.ihtm.bg.ac.rs/handle/123456789/3290]