Key role of quinone in the mechanism of respiratory complex I

Complex I is the first and the largest enzyme of respiratory chains in bacteria and mitochondria. The mechanism which couples spatially separated transfer of electrons to proton translocation in complex I is not known. Here we report five crystal structures of T. thermophilus enzyme in complex with NADH or quinone-like compounds. We also determined cryo-EM structures of major and minor native states of the complex, differing in the position of the peripheral arm. Crystal structures show that binding of quinone-like compounds (but not of NADH) leads to a related global conformational change, accompanied by local re-arrangements propagating from the quinone site to the nearest proton channel. Normal mode and molecular dynamics analyses indicate that these are likely to represent the first steps in the proton translocation mechanism. Our results suggest that quinone binding and chemistry play a key role in the coupling mechanism of complex I

Atomistic determinants of co-enzyme Q reduction at the Q(i)-site of the cytochrome bc(1) complex

The cytochrome (cyt) bc(1) complex is an integral component of the respiratory electron transfer chain sustaining the energy needs of organisms ranging from humans to bacteria. Due to its ubiquitous role in the energy metabolism, both the oxidation and reduction of the enzyme's substrate co-enzyme Q has been studied vigorously. Here, this vast amount of data is reassessed after probing the substrate reduction steps at the Q(i)-site of the cyt bc(1) complex of Rhodobacter capsulatus using atomistic molecular dynamics simulations. The simulations suggest that the Lys251 side chain could rotate into the Q(i)-site to facilitate binding of half-protonated semiquinone - a reaction intermediate that is potentially formed during substrate reduction. At this bent pose, the Lys251 forms a salt bridge with the Asp252, thus making direct proton transfer possible. In the neutral state, the lysine side chain stays close to the conserved binding location of cardiolipin (CL). This back-and-forth motion between the CL and Asp252 indicates that Lys251 functions as a proton shuttle controlled by pH-dependent negative feedback. The CL/K/D switching, which represents a refinement to the previously described CL/K pathway, fine-tunes the proton transfer process. Lastly, the simulation data was used to formulate a mechanism for reducing the substrate at the Q(i)-site.Peer reviewe

Cryo-EM structure of the fully assembled elongator complex

Transfer RNA (tRNA) molecules are essential to decode messenger RNA codons during protein synthesis. All known tRNAs are heavily modified at multiple positions through post-transcriptional addition of chemical groups. Modifications in the tRNA anticodons are directly influencing ribosome decoding and dynamics during translation elongation and are crucial for maintaining proteome integrity. In eukaryotes, wobble uridines are modified by Elongator, a large and highly conserved macromolecular complex. Elongator consists of two subcomplexes, namely Elp123 containing the enzymatically active Elp3 subunit and the associated Elp456 hetero-hexamer. The structure of the fully assembled complex and the function of the Elp456 subcomplex have remained elusive. Here, we show the cryo-electron microscopy structure of yeast Elongator at an overall resolution of 4.3 Å. We validate the obtained structure by complementary mutational analyses in vitro and in vivo. In addition, we determined various structures of the murine Elongator complex, including the fully assembled mouse Elongator complex at 5.9 Å resolution. Our results confirm the structural conservation of Elongator and its intermediates among eukaryotes. Furthermore, we complement our analyses with the biochemical characterization of the assembled human Elongator. Our results provide the molecular basis for the assembly of Elongator and its tRNA modification activity in eukaryotes

Arithmetical properties of certain sums including Legendre symbols

W pracy autor bada sumy symboli Legendre'a z wielomianów stopnia 2.In this paper author studies sums of Legendre symbols of polynomial with degree 2

Determination of volatile organic compounds in materials from polystyrene intended for contact with food: comparison of HS-GS/MS and SPME-GC/MS techniques

ABSTRACT Background. Plastic materials intended for contact with food sometimes exhibit unfavorable organoleptic properties which is related to the presence of volatile organic compounds. These substances not only worsen organoleptic properties, but can be very harmful to humans health. For the sake of consumer safety, it is necessary to control such materials for the content of substances harmful to health, as well as the possibility of their migration to food. Therefore, there is a need to have an appropriate and verified analytical method that could be used in the routine analysis of volatile organic substances present in food contact materials. Objective. In this study, the possibilities of the application of HS-GC/MS and SPME-GC/MS analytical techniques for analyses of volatile organic compounds present in polystyrene food contact materials, demonstrating disadvantageous organoleptic properties were evaluated. Materials and methods. On the basis of sensory tests four types of food contact materials (plastic dishes) were selected for the study. The analytical measurement was performed by HS-GC/MS and SPME-GC/MS techniques parallel. Results. In quality examinations of samples the aliphatic and aromatic, saturated and unsaturated hydrocarbons, as well as other compounds, e.g. ketones contamination was estimated. For all the samples a quantitative analysis of the content of styrene, ethylbenzene and cyclohexane was carried out. Additionally, the optimization of SPME analysis parameters was carried out. It was assumed that the optimal SPME extraction conditions for this purposes are: extraction time of 15 - 30 min, extraction temperature of 80°C, CAR/PDMS fibre. Conclusions. The results of this study indicated that from two chosen analytical methods, definitely HS-GC/MS technique was more universal, as well as more comfortable and faster. Sometimes, however additional studies should be undertaken and then it is recommended to use the SPME-GC/MS technique optimized for our purposes

Key role of water in proton transfer at the Q_{o}-site of the cytochrome bc_{1} complex predicted by atomistic molecular dynamics simulations

AbstractCytochrome (cyt) bc1 complex, which is an integral part of the respiratory chain and related energy-conserving systems, has two quinone-binding cavities (Qo- and Qi-sites), where the substrate participates in electron and proton transfer. Due to its complexity, many of the mechanistic details of the cyt bc1 function have remained unclear especially regarding the substrate binding at the Qo-site. In this work we address this issue by performing extensive atomistic molecular dynamics simulations with the cyt bc1 complex of Rhodobacter capsulatus embedded in a lipid bilayer. Based on the simulations we are able to show the atom-level binding modes of two substrate forms: quinol (QH2) and quinone (Q). The QH2 binding at the Qo-site involves a coordinated water arrangement that produces an exceptionally close and stable interaction between the cyt b and iron sulfur protein subunits. In this arrangement water molecules are positioned suitably in relation to the hydroxyls of the QH2 ring to act as the primary acceptors of protons detaching from the oxidized substrate. In contrast, water does not have a similar role in the Q binding at the Qo-site. Moreover, the coordinated water molecule is also a prime candidate to act as a structural element, gating for short-circuit suppression at the Qo-site