8,189 research outputs found

    Membrane systems with limited parallelism

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    Membrane computing is an emerging research field that belongs to the more general area of molecular computing, which deals with computational models inspired from bio-molecular processes. Membrane computing aims at defining models, called membrane systems or P systems, which abstract the functioning and structure of the cell. A membrane system consists of a hierarchical arrangement of membranes delimiting regions, which represent various compartments of a cell, and with each region containing bio-chemical elements of various types and having associated evolution rules, which represent bio-chemical processes taking place inside the cell. This work is a continuation of the investigations aiming to bridge membrane computing (where in a compartmental cell-like structure the chemicals to evolve are placed in compartments defined by membranes) and brane calculi (where one considers again a compartmental cell-like structure with the chemicals/proteins placed on the membranes themselves). We use objects both in compartments and on membranes (the latter are called proteins), with the objects from membranes evolving under the control of the proteins. Several possibilities are considered (objects only moved across membranes or also changed during this operation, with the proteins only assisting the move/change or also changing themselves). Somewhat expected, computational universality is obtained for several combinations of such possibilities. We also present a method for solving the NP-complete SAT problem using P systems with proteins on membranes. The SAT problem is solved in O(nm) time, where n is the number of boolean variables and m is the number of clauses for an instance written in conjunctive normal form. Thus, we can say that the solution for each given instance is obtained in linear time. We succeeded in solving SAT by a uniform construction of a deterministic P system which uses rules involving objects in regions, proteins on membranes, and membrane division. Then, we investigate the computational power of P systems with proteins on membranes in some particular cases: when only one protein is placed on a membrane, when the systems have a minimal number of rules, when the computation evolves in accepting or computing mode, etc. This dissertation introduces also another new variant of membrane systems that uses context-free rewriting rules for the evolution of objects placed inside compartments of a cell, and symport rules for communication between membranes. The strings circulate across membranes depending on their membership to regular languages given by means of regular expressions. We prove that these rewriting-symport P systems generate all recursively enumerable languages. We investigate the computational power of these newly introduced P systems for three particular forms of the regular expressions that are used by the symport rules. A characterization of ET0L languages is obtained in this context

    Computational studies of biomembrane systems: Theoretical considerations, simulation models, and applications

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    This chapter summarizes several approaches combining theory, simulation and experiment that aim for a better understanding of phenomena in lipid bilayers and membrane protein systems, covering topics such as lipid rafts, membrane mediated interactions, attraction between transmembrane proteins, and aggregation in biomembranes leading to large superstructures such as the light harvesting complex of green plants. After a general overview of theoretical considerations and continuum theory of lipid membranes we introduce different options for simulations of biomembrane systems, addressing questions such as: What can be learned from generic models? When is it expedient to go beyond them? And what are the merits and challenges for systematic coarse graining and quasi-atomistic coarse grained models that ensure a certain chemical specificity

    Logarithmic SAT Solution with Membrane Computing

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    P systems have been known to provide efficient polynomial (often linear) deterministic solutions to hard problems. In particular, cP systems have been shown to provide very crisp and efficient solutions to such problems, which are typically linear with small coefficients. Building on a recent result by Henderson et al., which solves SAT in square-root-sublinear time, this paper proposes an orders-of-magnitude-faster solution, running in logarithmic time, and using a small fixed-sized alphabet and ruleset (25 rules). To the best of our knowledge, this is the fastest deterministic solution across all extant P system variants. Like all other cP solutions, it is a complete solution that is not a member of a uniform family (and thus does not require any preprocessing). Consequently, according to another reduction result by Henderson et al., cP systems can also solve k-colouring and several other NP-complete problems in logarithmic time

    Polarizationless P Systems with Active Membranes: Computational Complexity Aspects

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    P systems with active membranes, in their classical definition, make use of noncooperative rules only. However, it is well known that in living cells, proteins interact among them yielding new products. Inspired by this biological phenomenon, the previous framework is reformulated in this paper, allowing cooperation in object evolution rules, while removing electrical charges associated with membranes. More precisely, minimal cooperation in object evolution rules is incorporated in polarizationless P systems with active membranes. In this paper, the term “minimal” means that the left-hand side of such rules consists of at most two symbols, and its length is greater than or equal to the corresponding right-hand side. The computational efficiency of this kind of P systems is studied by providing a uniform polynomial-time solution to SAT problem in such manner that only division rules for elementary membranes are used and dissolution rules are forbidden. Bearing in mind that only tractable problems can be efficiently solved by families of polarizationless P systems with active membranes and without dissolution rules, passing from non-cooperation to minimal cooperation in object evolution rules amounts passing from non-efficiency to efficiency in this framework. This frontier of efficiency provides, as any other borderline does, a possible way to address the P versus NP problem.National Natural Science Foundation of China No. 61033003National Natural Science Foundation of China No. 6132010600

    Minimal cooperation in polarizationless P systems with active membranes

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    P systems with active membranes is a well developed framework in the eld of Membrane Computing. Using evolution, communication, dissolution and division rules, we know that some kinds of problems can be solved by those systems, but taking into account which ingredients are used. All these rules are inspired by the behavior of living cells, who \compute" with their proteins in order to obtain energy, create components, send information to other cells, kill themselves (in a process called apoptosis), and so on. But there are other behaviors not captured in this framework. As mitosis is simulated by division rules (for elementary and non-elementary membranes), meiosis, that is, membrane ssion inspiration is captured in separation rules. It di ers from the rst in the sense of duplication of the objects (that is, in division rules, we duplicate the objects not involved in the rule, meanwhile in separation rules we divide the content of the original membrane into the new membranes created). Evolution rules simulate the transformation of components in membranes, but it is well known that elements interact with another ones in order to obtain new components. Cooperation in evolution rules is considered. More speci cally, minimal cooperation (in the sense that only two objects can interact in order to create one or two objects

    Morphological and Photoelectrochemical Characterization of Membrane Reconstituted Photosystem I (PSI)

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    The robust structural and photoactive electrochemical properties of Photosystem I (PSI), a transmembrane photosynthetic protein complex, make it an ideal candidate for incorporation into solid state bioelectronic or hybrid photovoltaic devices. However, the first step towards the successful fabrication of such devices requires systematic assembly of oriented and functional PSI onto desired bio-abio interfaces via suitable protein scaffoldings. Hence, this dissertation focuses on utilizing the cyanobacterial PSI for integration into organic/inorganic interfaces that mediate photo-electrochemical energy conversions for electricity and/or solar fuel production. To this end, in this study the effect of systematic incorporation of PSI complexes into synthetic membrane-bound structures that mimic the natural thylakoid membrane housing of PSI quantifies via its performance and photocurrent response is demonstrated. Therefore, the surfactant-induced membrane solubilization of three phospholipids, namely DPhPC (1,2-diphytanoyl-sn-glycero-3-phosphocholine), DPPG (1,2-dipalmitoyl-sn-glycero-3-phospho-(1\u27-rac-glycerol)), and DPhPG (1,2-diphytanoyl-sn-glycero-3-phospho-(1\u27-rac-glycerol)) with the motivation of creating biomimetic reconstructs of PSI reconstitution in these liposomes are studied via isothermal titration calorimetry, turbidity measurements, dynamic light scattering and cryo-transmission electron microscopy imaging. The results indicate the typical three-stage solubilization process during lamellar-to-micellar transitions for liposomes is dictated by the critical detergent/phospholipid ratios. Considering that most successful protein incorporation occurs during the second stage of solublization, these studies set the backdrop for ideal concentration ratios for successful protein insertion in this stage. Furthermore, a facile yet elegant method for incorporation of PSI trimeric complexes into DPhPG bilayer membranes is introduced. The efficacy of this method is demonstrated via absorption and fluorescence spectroscopy measurements as well as direct visualization using atomic force microscopy. This study also provides direct evidence that PSI confinements in synthetic lipid scaffolds can be used for tuning the photoexcitation characteristics of PSI. Finally, detailed chronoamperometry measurements were conducted on PSI-proteoliposomes made from PSI incorporated within biomimetic membrane scaffolds and supported on suitable SAM substrates to investigate the enhancement in photocurrent responses arising from such confinement. The significant observation here is that the photo currents generated from PSI complexes under liposome confinements produce photocurrents four times higher than that produced from dense monolayer of individual PSI on SAM substrates using an equivalent concentration of PSI

    Structural and functional characterization of PorA and PorH : the two major porins from Corynebacterium glutamicum

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    PorA (5 kDa) et PorH (7kDa) sont les deux protéines membranaires majeures de la membrane externe de Corynebacterium glutamicum qui appartient au groupe supragénérique des bactéries Gram-positives contenant plusieurs agents pathogÚnes i.e. Mycobacterium tuberculosis, M. leprae et C. diphtheriae. Les deux protéines forment des canaux ioniques hétéromériques et présentent la particularité d'avoir une modification post-traductionnelle : estérification par l'acide mycolique. Les deux protéines ont été produites dans deux systÚmes d'expression : chez C. glutamicum et dans un systÚme acellulaire utilisant des extraits de E. coli. La présence ou l'absence de modification post-traductionnelle, sur les protéines produites in vivo et in vitro a été caractérisée par spectrométrie de masse MALDI-TOF. Les spectres de dichroïsme cellulaire et de RMN de PorA et PorH uniformément marquées et solubilisées en micelles de LDAO sont caractéristiques de protéines mono-disperses, partiellement structurées, et de qualité compatible avec une détermination de structure par RMN. Le test fonctionnel des protéines, par mesure de conductivité ionique aprÚs reconstitution dans une membrane lipidique (technique dite de BLM pour " black lipid membrane ") a montré que a) la modification post-traductionnelle de PorA par un acide mycolique est essentielle (contrairement à celle de PorH) b) la présence simultanée de PorA et de PorH est requise pour la formation d'un canal ionique voltage dépendant typique d'une porine. Afin de mieux comprendre l'importance de l'acide mycolique pour l'activité canal ionique, le complexe protéique PorA-PorH a été reconstitué dans son environnement naturel. Les principaux lipides de la membrane externe du C. glutamicum [Tréhalose dimycolate (TDM), tréhalose monomycolate (TMM) et cardiolipide (CL)] ont été extraits et purifiés par chromatographie d'adsorption et échange d'ions, sous forme protonée et sous forme perdeutériée. AprÚs formation de protéoliposomes les propriétés membranaires de TDM seul ou en mélange avec CL ont été étudiées RMN du deutérium, diffusion dynamique de la lumiÚre et microscopie électronique. L'insertion de PorA et PorH dans des vésicules de TDM a permis de mettre en évidence la reconstitution de l'hétéro-oligomÚre (contrairement aux micelles de LDAO). Ceci ouvre la voie à la détermination de structure 3D du complexe PorA-PorH fonctionnel par RMN solide et/ou liquide.PorA (5 kDa) and PorH (7 kDa) are two major membrane proteins from the outer membrane of Corynebacterium glutamicum which belongs to the suprageneric group of Gram-positive bacteria containing number of human pathogens such as Mycobacterium tuberculosis, M. leprae and C. diphtheriae. Both PorA and PorH have been shown to form heteromeric ion channels and to be post-translationally modified by mycolic acids (a-alkyl, beta-hydroxy fatty acids). Both proteins were produced in their natural host with mycolic acid modification, as well as in E. coli based continuous exchange cell-free expression system and thus devoid of mycolic acid modification. The presence or absence of mycolic acid modification on in vivo and in vitro expressed proteins was confirmed by MALDI-TOF mass spectrometry. CD and NMR spectra of 15N/13C uniformly labeled PorA and PorH solubilized in LDAO micelles indicated mono-dispersed and partially folded proteins, compatible with structure determination by NMR. However, functional assays (via black lipid membrane ion-channel conductance measurements) confirmed that a complex associating both proteins is required for function and that the mycolic acid modification on PorA (but not PorH), is an absolute requirement for the formation of a voltage dependent ion-channel. To understand further the importance of covalent or non-covalent interaction of their natural lipid environment on the complex formation, the major C. glutamicum outer membrane lipids [Trehalose dimycolate (TDM), Trehalose monomycolate (TMM) and Cardiolipin (CL)] were purified using adsorption and ion exchange chromatography, both in protonated and perdeuterated form. Prior to proteoliposome reconstitution, the membrane forming properties of TDM alone or in mixture with CL were studied by 2H-NMR, Dynamic Light Scattering and Electron Microscopy. Furthermore, after in vitro reconstitution of PorA and PorH in TDM vesicles (and not in LDAO micelles or DMPC vesicles), evidence for the formation of the hetero oligomeric complex was observed. The 3D structure determination, by liquid and/or solid state NMR, of a functional PorA-PorH complex in its natural lipid environment is now feasible

    Towards an understanding of the self-assembly and applications of phospholipid-containing nanoparticles

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    Polymer-stabilized phospholipid nanodiscs are nanoscale, discoidal assemblies containing a central core of lipid bilayer stabilized in aqueous solution by an amphipathic copolymer belt. Recently, nanodiscs have been applied to the extraction of membrane proteins directly from cellular membranes. However, the rapid adoption of nanodisc technology for biomolecular studies has outpaced the communities understanding of the fundamental properties of nanodisc-forming polymers which influence nanodisc self-assembly, as well as the inherent properties of the nanodiscs themselves. In this study, the thermodynamics of nanodisc self-assembly have been probed in order to gain insights into the structural properties of existing and novel polymers which affect the self-assembly process. Subsequently, their application to solubilisation of biological membranes has been investigated, overcoming limitations of the current technology. Following these studies, investigations into the interaction of nanodiscs with pre-existing membranes at interfaces were performed, revealing how lipid exchange kinetics vary between nanodisc types. Furthermore, polymer-stabilised nanodiscs were observed to adsorb to lipid bilayers, enabling future surface-based studies of nanodisc-encapsulated membrane proteins. Finally, polymer-stabilized nanodiscs were shown to provide a soluble membrane surface for investigation of protein-lipid interactions. This has revealed a membrane-induced fibrilization of the bacterial lipoprotein YraP, providing clues as to its currently unknown function

    Optimization of Brownian ratchets for the manipulation of charged components within supported lipid bilayers

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    In probability theory, there is a counter-intuitive result that it is possible to construct a winning strategy from two individually losing (or at most breaking-even) "games" by alternating between them. The work presented here demonstrates the application of this principle to supported lipid bilayers (SLBs) in order to create directed motion of charged lipid components in the membrane, which was achieved through the use of "Brownian ratchets" in patterned SLBs. Both a finite element analysis model and an experimental setup have been used to investigate the role of key parameters for the operation of these ratchets: (1) the asymmetry of the ratchet teeth and (2) the relation of the ratchet height to the period of the applied electric field. Importantly, we find that the efficiency of the ratchet for a given charged species is dependent on the diffusion coefficient. This opens the possibility for separation of membrane species according to their size or viscous drag coefficient within the membrane

    New synthetic strategies to tethered bilayer lipid membranes

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    Inhaltszusammenfassung in der Originalsprache des Dokuments (englisch) Tethered bilayer lipid membranes provide an efficient, stable and versatile platform for the investigation of integrated membrane proteins. However, the incorporation of large proteins, as well as of proteins with a large submembrane part is still a very critical issue and therefore, further optimisation of the system is necessary. The central element of a tBLM is a lipid bilayer. Its proximal leaflet is, at least to some extend, covalently attached to a solid support via a spacer group. The anchor lipid consists of three distinct parts, a lipid headgroup, a spacer group and an anchor. All parts together influence the final bilayer properties. In the frame of this work, the synthesis of new thiolipids for tBLMs on gold has been investigated. The aim was to obtain molecules with longer spacers in order to increase the submembrane space. The systems obtained have been characterized using SPR and EIS. The results obtained during this study are multiple. First, the synthesis of a previously synthesized architecture was successfully scaled up in an industrial lab using a new synthetic approach. The synthesis of large amounts is now feasible. Then, the synthesis of the new thiolipids was carried out taking into account the following requirements: the increase of the submembrane space by having longer ethyleneglycol spacers, the attachment of the molecules to a gold substrate via a thiol bond, and the tunability of the lateral mobility by changing the lipid headgroup. Three different synthetic strategies have been investigated. The polymeric approach did not prove to be successful, merely because of the broad molecular weight distribution. The synthesis of heterofunctionally protected oligoethyleneglycols allowed to obtain ethyleneglycol moieties with 6 and 8 units, but the tedious purification steps gave very low yields. Finally, the block by block synthesis using ethyleneglycol precursors proved to be an efficient and fast method to synthesize the target molecules. Indeed, these were obtained with very high yields, and the separation was very efficient. A whole family of new compounds was obtained, having 6, 8 and 14 ethyleneglycol units and with mono- or diphytanyl lipid headgroups. This new pathway is a very promising synthetic strategy that can be used further in the development of new compounds of the tether system. The formation of bilayers was investigated for the different thiolipids mainly by using EIS. The electrical properties of a bilayer define the quality of the membrane and allow the study of the functionality of proteins embedded in such a system. Despite multiple trials to improve the system using self assembly, Langmuir Blodgett transfer, and detergent mixed vesicles, the new polymer thiolipids did not show as high electrical properties as tBLMs reported in the literature. Nevertheless, it was possible to show that a bilayer could be obtained for the different spacer lengths. These bilayers could be formed using self assembly for the first monolayer, and two different methods for bilayer formation, namely vesicle fusion and solvent exchange. We could furthermore show functional incorporation of the ion carrier valinomycin: the selective transport of K+ ions could be demonstrated. For DPHL, it was even possible to show the functional incorporation of the ion channel gramicidin. The influence of the spacer length is translated into an increase of the spacer capacitance, which could correspond to an increase in the capacity of charge accumulation in the submembrane space. The different systems need to be further optimised to improve the electrical properties of the bilayer. Moreover, the incorporation of larger proteins, and proteins bearing submembrane parts needs to be investigated. -------------------------------------------------------------------------------- Inhaltszusammenfassung in einer weiteren Sprache (deutsch) FestkörperunterstĂŒtzte, verankerte Lipid Doppelschichten (tBLM) bieten eine effiziente, stabile und vielseitige Basis fĂŒr die Untersuchung von inkorporierten Membranproteinen. Der Einbau von grossen Proteinen, sowie Proteinen mit submembranen DomĂ€nen ist allerdings immer noch problematisch, und deswegen ist eine weitere Optimierung des Systems notwendig. Das zentrale Element einer tBLM ist eine Lipid Doppelschicht. Die untere Monolage ist mit Hilfe einer Abstandshaltergruppe kovalent an eine OberflĂ€che gebunden. Das Ankerlipid besteht aus drei Teilen, einem Lipidkopf, einem Abstandshalter und einer Ankergruppe. Alle Teile beeinflussen die Eigenschaften der Doppelschicht. Im Rahmen dieser Arbeit wurde die Synthese von neuen Thiolipiden fĂŒr tBLMs auf einer GoldoberflĂ€che mit lĂ€ngeren Abstandshaltern zur Vergrösserung des Reservoirs unter der Membran untersucht. Die erhaltenen Systeme wurden mit SPR und EIS charakterisiert. Die Ergebnisse dieser Untersuchung sind vielfĂ€ltig. FĂŒr die Synthese eines bekannten Thiolipids wurde ein Syntheseweg fĂŒr IndustriemassstĂ€be entwickelt und optimiert. Die Synthese grösserer Mengen ist jetzt möglich. Neue Thiolipide wurde unter BerĂŒcksichtigung folgender Anforderungen geplant und synthetisiert: die Vergrösserung des Abstands zwischen Membran und Substrat mittels lĂ€ngerer Ethylenglykol-Abstandshalter; die Verankerung der MolekĂŒle auf einer OberflĂ€che mit Hilfe von Thiolbindungen; die VerĂ€nderung und Anpassung der lateralen Beweglichkeit der Membran durch verschiedene Lipidköpfe. Drei verschiedene Strategien wurden untersucht. Der Ansatz ĂŒber Polymerisation war nicht erfolgreich, hauptsĂ€chlich wegen der hohen PolydispersitĂ€t der erhaltenen MolekĂŒle. Die Synthese von heterofunktionalisierten Oligoethylenglykolen ermöglichte die Gewinnung von Ethylenglykolketten mit 6 und 8 Einheiten. Die aufwendigen Reinigungs- und Trennungsschritte erlaubten nur eine sehr niedrige Ausbeute. Die „bloc by bloc“ Synthese mit geschĂŒtzten Ethylenglykolgruppen war eine erfolgreiche und schnelle Methode zur Synthese der Zielverbindungen. Die Trennung war sehr effizient und die Verbindungen wurden mit sehr hohen Ausbeuten erhalten. Eine ganze Serie von neuen Thiolipiden, mit 6, 8 und 14 Ethylenglykol-Einheiten mit mono- oder diphytanyl Kopfgruppen konnte erhalten werden. Dieser neue Syntheseweg ist eine vielversprechende Strategie, die fĂŒr die Entwicklung neuer tBLMs Bestandteile. Die verschiedenen Synthetisierten Thiolipide wurden als Basis fĂŒr Lipid Doppelschichten verwendet. Die Charakterisierung der Systeme erfolgte vor allem per Impedanzspektroskopie. Die elektrischen Eigenschaften einer Doppelschicht sind ein wichtiges Kriterium fĂŒr die QualitĂ€t der Membran und erlauben die Untersuchung der FunktionalitĂ€t von eingebauten Proteinen. Obwohl viele Versuche zur Verbesserung des Systems z.B. mittels self assembly, LB transfer und Vesikeln gemischt mit Detergenz unternommen wurden, zeigen die Polymerthiolipide keine so guten elektrischen Eigenschaften wie in der Literatur beschriebene tBLMs. Es war hingegen möglich zu zeigen, dass sich aus allen hier diskutierten Verbindungen Membranen herstellen lassen. Diese Doppelschichten wurden durch Selbstorganisation des ersten Monolayers gebildet, und zwei verschiedene Methoden wurden fĂŒr die Doppelschichtbildung benutzt, Vesikelfusion und schneller Lösungsmittelwechsel. In den meisten FĂ€llen war der Einbau des Ionen-Transporters Valinomycin erfolgreich. Der selektive Transport von K+ Ionen wurde an diesem Beispiel gezeigt. FĂŒr DPHL konnte der Ionenkanal Gramicidin funktionell eingebaut werden. Die Auswirkung der AbstandslĂ€nge bewirkt eine Zunahme der AbstandskapazitĂ€t, die zu einer Erhöhung der Ladungsakkumulation im submembranen Teil der Membranarchitektur fĂŒhrt. Die verschiedenen Systeme mĂŒssen weiter optimiert werden, um die elektrischen Eigenschaften der Doppelschicht zu verbessern. Der nĂ€chste Schritt ist der Einbau von grösseren Proteinen mit submembranen DomĂ€nen
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