Browsing by Subject "CRYSTAL-STRUCTURE"

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  • Blum, Markus; Gebhardt, Jacqueline; Papendick, Marius; Schlindwein, Simon H.; Nieger, Martin; Gudat, Dietrich (2018)
    Two N-heterocyclic phosphines with exocyclic SCN substituents were synthesised via metathesis of chlorophosphine precursors with KSCN and fully characterised. The crystallographic studies reveal that the products exhibit pronounced structural differences. The thiocyanato unit binds in one case via the nitrogen atom to yield a molecular structure with a slightly elongated P-N single bond and, in the other case, via the sulfur atom to form a structure that is best described as an ion pair and forms a one-dimensional coordination polymer in the crystal. DFT calculations suggest that the P-N and P center dot center dot center dot S interactions can be described as covalent and dative bonds, respectively, and that the structural differences correlate with the different cation stabilities of the individual phosphenium cation fragments.
  • Haapanen, Outi; Reidelbach, Marco; Sharma, Vivek (2020)
    Respiratory complex I (NADH:quinone oxidoreductase) plays a central role in generating the proton electrochemical gradient in mitochondrial and bacterial membranes, which is needed to generate ATP. Several high-resolution structures of complex I have been determined, revealing its intricate architecture and complementing the biochemical and biophysical studies. However, the molecular mechanism of long-range coupling between ubiquinone (Q) reduction and proton pumping is not known. Computer simulations have been applied to decipher the dynamics of Q molecule in the similar to 30 angstrom long Q tunnel. In this short report, we discuss the binding and dynamics of Q at computationally predicted Q binding sites, many of which are supported by structural data on complex I. We suggest that the binding of Q at these sites is coupled to proton pumping by means of conformational rearrangements in the conserved loops of core subunits.
  • Mishra, Arjun K.; Megta, Abhin Kumar; Palva, Airi; von Ossowski, Ingemar; Krishnan, Vengadesan (2017)
    SpaE is the predicted basal pilin subunit in the sortase-dependent SpaFED pilus from the gut-adapted and commensal Lactobacillus rhamnosus GG. Thus far, structural characterization of the cell-wall-anchoring basal pilins has remained difficult and has been limited to only a few examples from pathogenic genera and species. To gain a further structural understanding of the molecular mechanisms that are involved in the anchoring and assembly of sortase-dependent pili in less harmful bacteria, L. rhamnosus GG SpaE for crystallization was produced by recombinant expression in Escherichia coli. Although several attempts to crystallize the SpaE protein were unsuccessful, trigonal crystals that diffracted to a resolution of 3.1 angstrom were eventually produced using PEG 3350 as a precipitant and high protein concentrations. Further optimization with a combination of additives led to the generation of SpaE crystals in an orthorhombic form that diffracted to a higher resolution of 1.5 angstrom. To expedite structure determination by SAD phasing, selenium-substituted (orthorhombic) SpaE crystals were grown and X-ray diffraction data were collected to 1.8 angstrom resolution.
  • Djikic, Teodora; Vucicevic, Jelica; Laurila, Jonne; Radi, Marco; Veljkovic, Nevena; Xhaard, Henri; Nikolic, Katarina M. (2020)
    Based on the finding that a central antihypertensive agent with high affinity for I1-type imidazoline receptors ? rilmenidine, shows cytotoxic effects on cultured cancer cell lines, it has been suggested that imidazoline receptors agonists might have a therapeutic potential in the cancer therapy. Nevertheless, potential rilmenidine side effects caused by activation of α-adrenoceptors, or other associated receptors and enzymes, might hinder its therapeutic benefits. Considering that human α-adrenoceptors belong to the rhodopsin-like class A of G-protein-coupled receptors (GPCRs) it is reasonable to assume that imidazolines might have the affinity for other receptors from the same class. Therefore, to investigate possible off-target effects of imidazoline ligands we have prepared a reverse docking protocol on class A GPCRs, using imidazoline ligands and their decoys. To verify our in silico results, three ligands with high scores and three ligands with low scores were tested for antagonistic activity on α2- adrenoceptors.
  • Hassan, Alaa A.; Aly, Ashraf A.; Mohamed, Nasr K.; El Shaieb, Kamal M.; Makhlouf, Maysa M.; Abdelhafez, El-Shimaa M. N.; Bräse, Stefan; Nieger, Martin; Dalby, Kevin N.; Kaoud, Tamer S. (2019)
    A large number of natural products containing the propellane scaffold have been reported to exhibit cytotoxicity against several cancers; however, their mechanism of action is still unknown. Anticancer drugs targeting DNA are mainly composed of small planar molecule/s that can interact with the DNA helix, causing DNA malfunction and cell death. The aim of this study was to design and synthesize propellane derivatives that can act as DNA intercalators and/or groove binders. The unique structure of the propellane derivatives and their ability to display planar ligands with numerous possible geometries, renders them potential starting points to design new drugs targeting DNA in cancer cells. New substituted furo-imidazo[3.3.3]propellanes were synthesized via the reaction of substituted alkenylidene-hydrazinecarbothioamides with 2-(1,3-dioxo-2,3-dihydr- 1H-2-ylidene) propanedinitrile in tetrahydrofuran at room temperature. The structures of the products were confirmed by a combination of elemental analysis, NMR, ESI-MS, IR and single crystal X-ray analysis. Interestingly, 5c, 5d and 5f showed an ability to interact with Calf Thymus DNA (CT-DNA). Their DNA-binding mode was investigated using a combination of absorption spectroscopy, DNA melting, viscosity, CD spectroscopy measurements, as well as competitive binding studies with several dyes. Their cytotoxicity was evaluated against the NCI-60 panel of cancer cell lines. 5c, 5d and 5f exhibited similar anti-proliferative activity against the A549 non-small cell lung cancer (NSCLC) cell line. Further mechanistic studies revealed their ability to induce DNA damage in the A549 cell line, as well as apoptosis, evidenced by elevated Annexin V expression, enhanced caspase 3/7 activation and PARP cleavage. In this study, we present the potential for designing novel propellanes to provoke cytotoxic activity, likely through DNA binding-induced DNA damage and apoptosis.
  • Baloban, Mikhail; Shcherbakova, Daria M.; Pletnev, Sergei; Pletnev, Vladimir Z.; Lagarias, J. Clark; Verkhusha, Vladislav (2017)
    Brighter near-infrared (NIR) fluorescent proteins (FPs) are required for multicolor microscopy and deep-tissue imaging. Here, we present structural and biochemical analyses of three monomeric, spectrally distinct phytochrome-based NIR FPs, termed miRFPs. The miRFPs are closely related and differ by only a few amino acids, which define their molecular brightness, brightness in mammalian cells, and spectral properties. We have identified the residues responsible for the spectral red-shift, revealed a new chromophore bound simultaneously to two cysteine residues in the PAS and GAF domains in blue-shifted NIR FPs, and uncovered the importance of amino acid residues in the N-terminus of NIR FPs for their molecular and cellular brightness. The novel chromophore covalently links the N-terminus of NIR FPs with their C-terminal GAF domain, forming a topologically closed knot in the structure, and also contributes to the increased brightness. Based on our studies, we suggest a strategy to develop spectrally distinct NIR FPs with enhanced brightness.
  • Väyrynen, Katja; Hatanpää, Timo; Mattinen, Miika; Heikkilä, Mikko; Mizohata, Kenichiro; Meinander, Kristoffer; Räisänen, Jyrki; Ritala, Mikko; Leskelä, Markku (2018)
    In this paper, we introduce a new Co precursor for the atomic layer deposition (ALD) of Co metal and other Co containing materials. CoCl2(TMEDA) (TMEDA = N,N,N ',N '-tetramethylethylenediamine) is a diamine adduct of cobalt(II) chloride that is inexpensive and easy to synthesize, making it an industrially viable precursor. Furthermore, CoCl2(TMEDA) shows good volatility at reasonably low temperatures and is thermally stable up to a temperature of, similar to 300 degrees C. We also present a full ALD study for the deposition of CoO thin films using CoCl2(TMEDA) and water as precursors. The process was investigated within a temperature range of 225-300 degrees C. Saturation of the film growth with respect to both precursor pulse lengths was verified. According to X-ray diffraction, the films were a mixture of hexagonal and cubic CoO. No reflections corresponding to Co3O4 were detected. The hexagonal phase is characteristic to nanomaterials only and is not seen in bulk CoO. The crystal structure of the films could be tuned by temperature, water pulse lengths, and type of substrate material. The films deposited at 275 degrees C exhibited 1:1 Co:O stoichiometry and very high purity. The CoO films could be reduced to Co metal at an exceptionally low temperature of 250 degrees C in 10% forming gas. Continuity of the reduced Co films was improved when the CoO film was deposited on TiN instead of native oxide terminated Si. The Co content of a 50 nm reduced metal film was as high as 95 at. %, with negligible amounts of oxygen and hydrogen.
  • Tailford, Louise E.; Owen, C. David; Walshaw, John; Crost, Emmanuelle H.; Hardy-Goddard, Jemma; Le Gall, Gwenaelle; de Vos, Willem M.; Taylor, Garry L.; Juge, Nathalie (2015)
    The gastrointestinal mucus layer is colonized by a dense community of microbes catabolizing dietary and host carbohydrates during their expansion in the gut. Alterations in mucosal carbohydrate availability impact on the composition of microbial species. Ruminococcus gnavus is a commensal anaerobe present in the gastrointestinal tract of > 90% of humans and overrepresented in inflammatory bowel diseases (IBD). Using a combination of genomics, enzymology and crystallography, we show that the mucin-degrader R. gnavus ATCC 29149 strain produces an intramolecular trans-sialidase (IT-sialidase) that cleaves off terminal alpha 2-3-linked sialic acid from glycoproteins, releasing 2,7-anhydro-Neu5Ac instead of sialic acid. Evidence of IT-sialidases in human metagenomes indicates that this enzyme occurs in healthy subjects but is more prevalent in IBD metagenomes. Our results uncover a previously unrecognized enzymatic activity in the gut microbiota, which may contribute to the adaptation of intestinal bacteria to the mucosal environment in health and disease.
  • Kaurola, Petri; Sharma, Vivek; Vonk, Amanda; Vattulainen, Ilpo; Rog, Tomasz (2016)
    Quinone and its analogues (Q) constitute an important class of compounds that perform key electron transfer reactions in oxidative- and photo-phosphorylation. In the inner membrane of mitochondria, ubiquinone molecules undergo continuous redox transitions enabling electron transfer between the respiratory complexes. In such a dynamic system undergoing continuous turnover for ATP synthesis, an uninterrupted supply of substrate molecules is absolutely necessary. In the current work, we have performed atomistic molecular dynamics simulations and free energy calculations to assess the structure, dynamics, and localization of quinone and its analogues in a lipid bilayer, whose composition mimics the one in the inner mitochondrial membrane. The results show that there is a strong tendency of both quinone and quinol molecules to localize in the vicinity of the lipids' acyl groups, right under the lipid head group region. Additionally, we observe a second location in the middle of the bilayer where quinone molecules tend to stabilize. Translocation of quinone through a lipid bilayer is very fast and occurs in 10-100 ns time scale, whereas the translocation of quinol is at least an order of magnitude slower. We suggest that this has important mechanistic implications given that the localization of Q ensures maximal occupancy of the Q-binding sites or Q-entry points in electron transport chain complexes, thereby maintaining an optimal turnover rate for ATP synthesis. (C) 2016 Elsevier B.V. All rights reserved.
  • Lewis, Samuel H.; Salmela, Heli; Obbard, Darren J. (2016)
    Genetic studies of Drosophila melanogaster have provided a paradigm for RNA interference (RNAi) in arthropods, in which the microRNA and antiviral pathways are each mediated by a single Argonaute (Ago1 and Ago2) and germline suppression of transposable elements is mediated by a trio of Piwi-subfamily Argonaute proteins (Ago3, Aub, and Piwi). Without a suitable evolutionary context, deviations from this can be interpreted as derived or idiosyncratic. Here we analyze the evolution of Argonaute genes across the genomes and transcriptomes of 86 Dipteran species, showing that variation in copy number can occur rapidly, and that there is constant flux in some RNAi mechanisms. The lability of the RNAi pathways is illustrated by the divergence of Aub and Piwi (182-156Ma), independent origins of multiple Piwi-family genes in Aedes mosquitoes (less than 25Ma), and the recent duplications of Ago2 and Ago3 in the tsetse fly Glossina morsitans. In each case the tissue specificity of these genes has altered, suggesting functional divergence or innovation, and consistent with the action of dynamic selection pressures across the Argonaute gene family. We find there are large differences in evolutionary rates and gene turnover between pathways, and that paralogs of Ago2, Ago3, and Piwi/Aub show contrasting rates of evolution after duplication. This suggests that Argonautes undergo frequent evolutionary expansions that facilitate functional divergence.
  • Suomivuori, Carl-Mikael; Gamiz-Hernandez, Ana P.; Sundholm, Dage; Kaila, Ville R. I. (2017)
    The conversion of light energy into ion gradients across biological membranes is one of the most fundamental reactions in primary biological energy transduction. Recently, the structure of the first light-activated Na+ pump, Krokinobacter eikastus rhodopsin 2 (KR2), was resolved at atomic resolution [Kato HE, et al. (2015) Nature 521: 48-53]. To elucidate its molecular mechanism for Na+ pumping, we perform here extensive classical and quantum molecular dynamics (MD) simulations of transient photocycle states. Our simulations show how the dynamics of key residues regulate water and ion access between the bulk and the buried light-triggered retinal site. We identify putative Na+ binding sites and show how protonation and conformational changes gate the ion through these sites toward the extracellular side. We further show by correlated ab initio quantum chemical calculations that the obtained putative photocycle intermediates are in close agreement with experimental transient optical spectroscopic data. The combined results of the ion translocation and gating mechanisms in KR2 may provide a basis for the rational design of novel light-driven ion pumps with optogenetic applications.
  • Galemou Yoga, Etienne; Parey, Kristian; Djurabekova, Amina; Haapanen, Outi; Siegmund, Karin; Zwicker, Klaus; Sharma, Vivek; Zickermann, Volker; Angerer, Heike (2020)
    Respiratory complex I catalyzes electron transfer from NADH to ubiquinone (Q) coupled to vectorial proton translocation across the inner mitochondrial membrane. Despite recent progress in structure determination of this very large membrane protein complex, the coupling mechanism is a matter of ongoing debate and the function of accessory subunits surrounding the canonical core subunits is essentially unknown. Concerted rearrangements within a cluster of conserved loops of central subunits NDUFS2 (beta 1-beta 2(S2) loop), ND1 (TMH5-6(ND1) loop) and ND3 (TMH1-2(ND3) loop) were suggested to be critical for its proton pumping mechanism. Here, we show that stabilization of the TMH1-2(ND3) loop by accessory subunit LYRM6 (NDUFA6) is pivotal for energy conversion by mitochondrial complex I. We determined the high-resolution structure of inactive mutant F89A(LYRM6) of eukaryotic complex I from the yeast Yarrowia lipolytica and found long-range structural changes affecting the entire loop cluster. In atomistic molecular dynamics simulations of the mutant, we observed conformational transitions in the loop cluster that disrupted a putative pathway for delivery of substrate protons required in Q redox chemistry. Our results elucidate in detail the essential role of accessory subunit LYRM6 for the function of eukaryotic complex I and offer clues on its redox-linked proton pumping mechanism. Respiratory complex I plays a key role in energy metabolism. Cryo-EM structure of a mutant accessory subunit LYRM6 from the yeast Yarrowia lipolytica and molecular dynamics simulations reveal conformational changes at the interface between LYRM6 and subunit ND3, propagated further into the complex. These findings offer insight into the mechanism of proton pumping by respiratory complex I.
  • Mönttinen, Heli A. M.; Ravantti, Janne J.; Poranen, Minna M. (2012)
  • Dilokpimol, Adiphol; Mäkelä, Miia Riitta; Varriale, Simona; Zhou, Miaomiao; Cerullo, Gabriella; Gidijala, Loknath; Hinkka, Harri Tapio; Brás, Joana L.A.; Jütten, Peter; Piechot, Alexander; Verhaert, Raymond; Hilden, Sari Kristiina; Faraco, Vincenza; de Vries, Ronald (2018)
    Feruloyl esterases (FAEs) are a diverse group of enzymes that specifically catalyze the hydrolysis of ester bonds between a hydroxycinnamic (e.g. ferulic) acid and plant poly- or oligosaccharides. FAEs as auxiliary enzymes significantly assist xylanolytic and pectinolytic enzymes in gaining access to their site of action during biomass saccharification for biofuel and biochemical production. A limited number of FAEs have been functionally characterized compared to over 1000 putative fungal FAEs that were recently predicted by similarity-based genome mining, which divided phylogenetically into different subfamilies (SFs). In this study, 27 putative and six characterized FAEs from both ascomycete and basidiomycete fungi were selected and heterologously expressed in Pichia pastoris and the recombinant proteins biochemically characterized to validate the previous genome mining and phylogenetical grouping and to expand the information on activity of fungal FAEs. As a result, 20 enzymes were shown to possess FAE activity, being active towards pNP-ferulate and/or methyl hydroxycinnamate substrates, and covering 11 subfamilies. Most of the new FAEs showed activities comparable to those of previously characterized fungal FAEs.
  • Choi, Jaeyoung; Kim, Ki-Tae; Jeon, Jongbum; Wu, Jiayao; Song, Hyeunjeong; Asiegbu, Fred O.; Lee, Yong-Hwan (2014)
  • Sunjuk, Mahmoud; Abu-Surrah, Adnan S.; Abu Safieh, Kayed A.; Qaroush, Abdussalam K.; Al-Qaisi, Feda'a M. (2017)
    The synthesis of new palladium(II) complexes of the type [Pd(A-N=C-ph-C=N-A) Cl-2] (4a-e) (A = cyclohexyl (a), 2-isoprpropyl (b), pyrenyl (c), naphthyl (d), and 2,6-diisopropyl (e)) is described. The isolated gamma-diimine ligands and their corresponding palladium(II) complexes were characterized by their physical properties, elemental analysis, H-1 NMR=, C-13 NMR, and infrared spectroscopy. The palladium(II) complexes (4a-e) were employed successfully as catalysts for atom transfer radical polymerization (ATRP) of methyl methacrylate (MMA) in the presence of ethyl-2-bromoisobutyrate (EBIB) as initiator at 90 degrees C. Polymerization with these catalyst systems afforded polymers with low molecular weight distribution (M-w/M-n) and syndio-rich atactic poly (MMA) with relatively higher [rr] diads. (C) 2013 Production and hosting by Elsevier B.V.
  • Wei, Gong-Hong; Badis, Gwenael; Berger, Michael F.; Kivioja, Teemu; Palin, Kimmo; Enge, Martin; Bonke, Martin; Jolma, Arttu; Varjosalo, Markku; Gehrke, Andrew R.; Yan, Jian; Talukder, Shaheynoor; Turunen, Mikko; Taipale, Mikko; Stunnenberg, Hendrik G.; Ukkonen, Esko; Hughes, Timothy R.; Bulyk, Martha L.; Taipale, Jussi (2010)
  • Polley, Anirban; Orlowski, Adam; Danne, Reinis; Gurtovenko, Andrey A.; de la Serna, Jorge Bernardino; Eggeling, Christian; Davis, Simon J.; Rog, Tomasz; Vattulainen, Ilpo (2017)
    Proteins embedded in the plasma membrane mediate interactions with the cell environment and play decisive roles in many signaling events. For cell-cell recognition molecules, it is highly likely that their structures and behavior have been optimized in ways that overcome the limitations of membrane tethering. In particular, the ligand binding regions of these proteins likely need to be maximally exposed. Here we show by means of atomistic simulations of membrane-bound CD2, a small cell adhesion receptor expressed by human T-cells and natural killer cells, that the presentation of its ectodomain is highly dependent on membrane lipids and receptor glycosylation acting in apparent unison. Detailed analysis shows that the underlying mechanism is based on electrostatic interactions complemented by steric interactions between glycans in the protein and the membrane surface. The findings are significant for understanding the factors that render membrane receptors accessible for binding and signaling.
  • Wannarit, Nanthawat; Pakawatchai, Chaveng; Mutikainen, Ilpo; Costa, Ramon; Moreira, Iberio de P. R.; Youngme, Sujittra; Illas, Francesc (2013)
  • Yetukuri, Laxman; Huopaniemi, Ilkka; Koivuniemi, Artturi; Maranghi, Marianna; Hiukka, Anne; Nygren, Heli; Kaski, Samuel; Taskinen, Marja-Riitta; Vattulainen, Ilpo; Jauhiainen, Matti; Oresic, Matej (2011)