Добірка наукової літератури з теми "Multi-protein assembly"

Abstract Binding of proteins to particular DNA sites across the genome is a primary determinant of specificity in genome maintenance and gene regulation. DNA-binding specificity is encoded at multiple levels, from the detailed biophysical interactions between proteins and DNA, to the assembly of multi-protein complexes. At each level, variation in the mechanisms used to achieve specificity has led to difficulties in constructing and applying simple models of DNA binding. We review the complexities in protein–DNA binding found at multiple levels and discuss how they confound the idea of simple recognition codes. We discuss the impact of new high-throughput technologies for the characterization of protein–DNA binding, and how these technologies are uncovering new complexities in protein–DNA recognition. Finally, we review the concept of multi-protein recognition codes in which new DNA-binding specificities are achieved by the assembly of multi-protein complexes.

Abstract The widespread occurrence of repetitive stretches of DNA in genomes of organisms across the tree of life imposes fundamental challenges for sequencing, genome assembly, and automated annotation of genes and proteins. This multi-level problem can lead to errors in genome and protein databases that are often not recognized or acknowledged. As a consequence, end users working with sequences with repetitive regions are faced with ‘ready-to-use’ deposited data whose trustworthiness is difficult to determine, let alone to quantify. Here, we provide a review of the problems associated with tandem repeat sequences that originate from different stages during the sequencing-assembly-annotation-deposition workflow, and that may proliferate in public database repositories affecting all downstream analyses. As a case study, we provide examples of the Atlantic cod genome, whose sequencing and assembly were hindered by a particularly high prevalence of tandem repeats. We complement this case study with examples from other species, where mis-annotations and sequencing errors have propagated into protein databases. With this review, we aim to raise the awareness level within the community of database users, and alert scientists working in the underlying workflow of database creation that the data they omit or improperly assemble may well contain important biological information valuable to others.

Many biological processes are governed by large assemblies of protein molecules. However, it is often very difficult to determine the three-dimensional structures of these assemblies using experimental biophysical techniques. Hence there is a need to develop computational approaches to fill this gap. This article presents an ant colony optimization approach to predict the structure of large multi-component protein complexes. Starting from pair-wise docking predictions, a multi-graph consisting of vertices representing the component proteins and edges representing candidate interactions is constructed. This allows the assembly problem to be expressed in terms of searching for a minimum weight spanning tree. However, because the problem remains highly combinatorial, the search space cannot be enumerated exhaustively and therefore heuristic optimisation techniques must be used. The utility of the ant colony based approach is demonstrated by re-assembling known protein complexes from the Protein Data Bank. The algorithm is able to identify near-native solutions for five of the six cases tested. This demonstrates that the ant colony approach provides a useful way to deal with the highly combinatorial multi-component protein assembly problem.

In the last three decades, many giant DNA viruses have been discovered. Giant viruses present a unique and essential research frontier for studies of self-assembly and regulation of supramolecular assemblies. The question on how these giant DNA viruses assemble thousands of proteins so accurately to form their protein shells, the capsids, remains largely unanswered. Revealing the mechanisms of giant virus assembly will help to discover the mysteries of many self-assembly biology problems. Paramecium bursaria Chlorella virus-1 (PBCV-1) is one of the most intensively studied giant viruses. Here, we implemented a multi-scale approach to investigate the interactions among PBCV-1 capsid building units called capsomers. Three binding modes with different strengths are found between capsomers around the relatively flat area of the virion surface at the icosahedral 2-fold axis. Furthermore, a capsomer structure manipulation package is developed to simulate the capsid assembly process. Using these tools, binding forces among capsomers were investigated and binding funnels were observed that were consistent with the final assembled capsid. In addition, total binding free energies of each binding mode were calculated. The results helped to explain previous experimental observations. Results and tools generated in this work established an initial computational approach to answer current unresolved questions regarding giant virus assembly mechanisms. Results will pave the way for studying more complicated process in other biomolecular structures.

Nuclear bodies (NBs) are structures that concentrate proteins, RNAs, and ribonucleoproteins that perform functions essential to gene expression. How NBs assemble is not well understood. We studied the Drosophila histone locus body (HLB), a NB that concentrates factors required for histone mRNA biosynthesis at the replication-dependent histone gene locus. We coupled biochemical analysis with confocal imaging of both fixed and live tissues to demonstrate that the Drosophila Multi Sex Combs (Mxc) protein contains multiple domains necessary for HLB assembly. An important feature of this assembly process is the self-interaction of Mxc via two conserved N-terminal domains: a LisH domain and a novel self-interaction facilitator (SIF) domain immediately downstream of the LisH domain. Molecular modeling suggests that the LisH and SIF domains directly interact, and mutation of either the LisH or the SIF domain severely impairs Mxc function in vivo, resulting in reduced histone mRNA accumulation. A region of Mxc between amino acids 721 and 1481 is also necessary for HLB assembly independent of the LisH and SIF domains. Finally, the C-terminal 195 amino acids of Mxc are required for recruiting FLASH, an essential histone mRNA-processing factor, to the HLB. We conclude that multiple domains of the Mxc protein promote HLB assembly in order to concentrate factors required for histone mRNA biosynthesis.

Yeast Dnm1p is a soluble, dynamin-related GTPase that assembles on the outer mitochondrial membrane at sites where organelle division occurs. Although these Dnm1p-containing complexes are thought to trigger constriction and fission, little is known about their composition and assembly, and molecules required for their membrane recruitment have not been isolated. Using a genetic approach, we identified two new genes in the fission pathway, FIS1 and FIS2. FIS1 encodes a novel, outer mitochondrial membrane protein with its amino terminus exposed to the cytoplasm. Fis1p is the first integral membrane protein shown to participate in a eukaryotic membrane fission event. In a related study (Tieu, Q., and J. Nunnari. 2000. J. Cell Biol. 151:353–365), it was shown that the FIS2 gene product (called Mdv1p) colocalizes with Dnm1p on mitochondria. Genetic and morphological evidence indicate that Fis1p, but not Mdv1p, function is required for the proper assembly and distribution of Dnm1p-containing fission complexes on mitochondrial tubules. We propose that mitochondrial fission in yeast is a multi-step process, and that membrane-bound Fis1p is required for the proper assembly, membrane distribution, and function of Dnm1p-containing complexes during fission.

Supramolecular assembly in vitro is a simple and effective way to produce multi-level biostructures to mimic the self-assembly of biomolecules in organisms. The study on peptide assembly behaviors would benefit a lot to understand what goes on in life, as well as in the construction of plenty of functional biomaterials that have potential applications in various fields. Since cellular microenvironments are crowded and contain various biomolecules, studying protein and peptide co-assembly is of great interest. Here, we introduced the co-assembly of 5-FAM-ELVFFAE-NH2 (EE-7) and (CY5)-KLVFFAK-NH2 (KK-7), which are sequences derived from the core of the amyloid β (Aβ) peptide, a key protein in Alzheimer’s diseases. Morphologic studies employing atomic force microscopy and scanning electron microscopy indicated that the co-assembled entities had a novel hydrangea-like microstructure, in contrast to micro-sheet structures formed from monocomponent EE-7 or KK-7, respectively. Fluorescence co-localization experiments confirmed that the hydrangealike microstructures were indeed made of both EE-7 and KK-7. We suggest that the formation of the hydrangea-like microstructures is driven by both the electrostatic and hydrophobic interactions between EE-7 and KK-7. A molecular mechanism has been provided to explain the formation of the hydrangea-like microstructures.

Electron transfer phenomena in proteins represent one of the most common types of biochemical reactions. They play a central role in energy conversion pathways in living cells, and are crucial components in respiration and photosynthesis. These complex biochemical reaction cascades consist of a series of proteins and protein complexes that couple a charge transfer to different forms of chemical energy. The efficiency and sophisticated optimisation of signal transfer in these natural redox chains has inspired engineering of artificial architectures mimicking essential properties of their natural analogues. Implementation of direct electron transfer (DET) in protein assemblies was a breakthrough in bioelectronics, providing a simple and efficient way for coupling biological recognition events to a signal transducer. DET avoids the use of redox mediators, reducing potential interferences and side reactions, as well as being more compatible with in vivo conditions. However, only a few haem proteins, including the redox protein cytochrome c (cyt.c), and blue copper enzymes show efficient DET on different kinds of electrodes. Previous investigations with cyt.c have mainly focused on heterogeneous electron transfer of monolayers of this protein on gold. An important advance was the fabrication of cyt.c multilayers by electrostatic layer-by-layer self-assembly. The ease of fabrication, the stability, and the controllable permeability of polyelectrolyte multilayers have made them particularly attractive for electroanalytical applications. With cyt.c and sulfonated polyaniline it was for the first time possible that fully electro-active multilayers of the redox protein could be prepared. This approach was extended to design an analytical signal chain based on multilayers of cyt.c and xanthine oxidase (XOD). The system does not need an external mediator but relies on an in situ generation of a mediating radical and thus allows a signal transfer from hypoxanthine via the substrate converting enzyme and cyt.c to the electrode. Another kind of a signal chain is based on assembling proteins in complexes on electrodes in such a way that a direct protein-protein electron transfer becomes feasible. This design does not need a redox mediator in analogy to natural protein communication. For this purpose, cyt.c and the enzyme bilirubin oxidase (BOD, EC 1.3.3.5) are co-immobilized in a self-assembled polyelectrolyte multilayer on gold electrodes. Although these two proteins are not natural reaction partners, the protein architecture facilitates an electron transfer from the electrode via multiple protein layers to molecular oxygen resulting in a significant catalytic reduction current. Finally, we describe a novel strategy for multi-protein layer-by-layer self-assembly combining cyt.c with an enzyme sulfite oxidase (SOx) without use of any additional polymer. Electrostatic interactions between these two proteins with rather separated pI values during the assembly process from a low ionic strength buffer were found sufficient for the layer-by-layer deposition of the both biomolecules. It is anticipated that the concepts described in this work will stimulate further progress in multilayer design of even more complex biomimetic signal cascades taking advantage of direct communication between proteins.
Elektronentransferphänomene in Proteinen stellen den häufigsten Typ biochemischer Reaktionen dar. Sie spielen eine zentrale Rolle bei der Energieumwandlung in der Zelle und sind entscheidende Komponenten in der Atmung und Photosynthese. Diese komplexen Kaskaden biochemischer Reaktionen setzen sich aus einer Reihe von Proteinen und Proteinkomplexen zusammen, die den Energietransfer an verschiedene Formen chemischer Energie koppeln. Die große Effektivität und Selektivität des Signaltransfers in diesen natürlichen Redoxketten war Vorbild für die Entwicklung künstlicher Architekturen, die die wesentlichen Eigenschaften ihrer natürlichen Analoga nachahmen. Die Implementierung des direkten Elektronentransfers (DET) von Proteinen mit Elektroden war ein Durchbruch im Bereich der Bioelektronik. Sie lieferte einen einfachen und effizienten Weg für das Koppeln biologischer Erkennungsereignisse an einen Signalumwandler. Durch den DET können Redoxmediatoren vermieden und damit potentielle Grenzflächen und Nebenreaktionen reduziert werden. Ebenso wird damit die Kompatibilität für in vivo Bedingungen erhöht. Jedoch zeigen nur einige Hämproteine wie das Redoxprotein Cytochrom c (Cyt c) und blaue Kupferproteine einen effizienten DET auf verschiedenen Elektrodentypen. Bisherige Untersuchungen mit Cyt c konzentrierten sich hauptsächlich auf den heterogenen Elektronentransfer von Monoschichten dieses Proteins auf Gold. Ein wichtiger Fortschritt war die Herstellung von Cyt c Multischichten durch die elektrostatische Layer-by-Layer-Technik. Die einfache Herstellung, die Stabilität sowie die kontrollierbaren Permeationseigenschaften von Polyelektrolyt-Multischichten machte sie besonders attraktiv für elektroanalytische Anwendungen. So gelang es auch zum ersten Mal vollständig elektroaktive Multischichten aus Cyt c und Polyanilinsulfonsäure zu präparieren. Dieser Ansatz wurde hier erweitert, um eine analytische Signalkette auf der Basis von Multischichten aus Cyt c und Xanthinoxidase zu entwerfen. Das System bedarf keinen externen Mediator, es hängt jedoch von der in situ Generierung eines vermittelnden Radikals ab und erlaubt daher einen Signaltransfer von Hypoxanthin über ein substratumwandelndes Enzym und Cyt c zur Elektrode. Eine andere Art von Signalketten basiert auf der Assemblierung von Proteinen in Komplexen auf Elektroden in solcher Art und Weise, daß ein direkter Protein-Protein-Elektronentransfer möglich wird. Dieser Ansatz benötigt keinen Redoxmediator in Analogie zu Beispielen aus dem biologischen Signaltransfer. Zu diesem Zweck werden Cyt c und das Enzym Bilirubinoxidase mit einem selbst-assemblierenden Polyelektrolyten auf einer Goldelektrode koimmobilisiert. Obwohl diese zwei Proteine keine natürlichen Reaktionspartner sind, unterstützt die Protein-Architektur einen Elektronentransfer von der Elektrode über mehrere Proteinschichten zu molekularem Sauerstoff und ergibt einen signifikanten katalytischen Reduktionsstrom. Schließlich wird eine neue Strategie beschrieben für eine Selbstassemblierung von Proteinen ohne zusätzlichen Polyelektrolyten - am Beispiel der Kombination von Cyt c mit Sulfitoxidase. Es stellte sich heraus, daß die elektrostatische Wechselwirkung zwischen diesen zwei Proteinen mit ziemlich weit voneinander entfernt liegenden pI-Werten während des Assemblierungsprozesses durch einen Puffer mit geringer Ionenstärke ausreicht um die beiden Biomoleküle nach dem Layer-by-Layer-Prinzip auf einer Elektrode abzuscheiden. Es wird erwartet, daß das entwickelte Konzept von Multiprotein-Assemblaten auf Elektroden weitere Fortschritte bei dem Entwurf von Multischichten und sogar noch komplexeren biomimetischen Signalkaskaden anregen wird und dabei der Vorteil der direkten Kommunikation zwischen Proteinen genutzt wird.

L'adsorption de protéine sur une surface artificielle solide est un phénomène fondamental qui détermine la réponse biologique d'un organisme vivant entrant dans n'importe quel matériel d'implant. Donc, l'adaptation de surfaces pour l'adsorption de protéine contrôlée est au coeur de beaucoup de champs de recherche d'aujourd'hui incluant la science de matériels et la biotechnologie. Dans ce contexte, les matériels sensibles de stimulus qui peuvent changer leurs propriétés comme une réponse à une petite monnaie dans leur environnement physicochimique attirent un grand intérêt comme ils permettent la création de surfaces avec des propriétés commutables pour le contrôle d'adsorption de protéine. Dans cette thèse, nous faisons un rapport sur la conception et l'élaboration de films minces sensibles de stimulus multi et de nanotubes. À cette fin, nous avons employé la couche-par-couche robuste et polyvalente…
Protein adsorption on a solid artificial surface is a fundamental phenomenon that determines the biological response of a living organism entering any implant material. Therefore, tailoring surfaces for controlled protein adsorption is at the heart of many of today's research fields including biotechnology and materials science. In this context, stimuli-responsive materials that are able to change their properties as a response to a small change in their physico-chemical environment are attracting a great interest as they allow the creation of surfaces with switchable properties for the control of protein adsorption. In this thesis, we report on the design and elaboration of multi stimuli-responsive thin films and nanotubes. For this purpose, we employed the robust and versatile layer-by-layer (LbL) assembly technique to incorporate block copolymers made of poly(acrylic acid) PAA and poly(N-isopropylacrylamide) PNIPAM with tunable and well-controlled block lengths. The combination of ellipsometry, quartz crystal microbalance with dissipation monitoring (QCM-D), surface plasmon resonance (SPR) and infrared data reveal the possibility to build up (PAH/PAA-b-PNIPAM)n multilayers. The stimuli-responsive properties of these LbL films were examined by monitoring the adsorption of proteins by means of QCM-D and fluorescence measurements, while varying (i) temperature, (ii) pH, (iii) ionic strength, or (iv) a combination of the above parameters. It appears that all these stimuli strongly influence the amount of adsorbed proteins. In short, these new PNIPAM block copolymer-based LbL coatings are easy to build on substrates of various nature and geometry (including nanoporous membranes)

Sequence alignment is a vital process in many biological applications such as Phylogenetic trees construction, DNA fragment assembly and structure/function prediction. Two kinds of alignment are pairwise alignment which align two sequences and Multiple Sequence alignment (MSA) that align sequences more than two. The accurate method of alignment is based on Dynamic Programming (DP) approach which suffering from increasing time exponentially with increasing the length and the number of the aligned sequences. Stochastic or meta-heuristics techniques speed up alignment algorithm but with near optimal alignment accuracy not as that of DP. Hence, This chapter aims to review the recent development of MSA using meta-heuristics algorithms. In addition, two recent techniques are focused in more deep: the first is Fragmented protein sequence alignment using two-layer particle swarm optimization (FTLPSO). The second is Multiple sequence alignment using multi-objective based bacterial foraging optimization algorithm (MO-BFO).

Sequence alignment is a vital process in many biological applications such as Phylogenetic trees construction, DNA fragment assembly and structure/function prediction. Two kinds of alignment are pairwise alignment which align two sequences and Multiple Sequence alignment (MSA) that align sequences more than two. The accurate method of alignment is based on Dynamic Programming (DP) approach which suffering from increasing time exponentially with increasing the length and the number of the aligned sequences. Stochastic or meta-heuristics techniques speed up alignment algorithm but with near optimal alignment accuracy not as that of DP. Hence, This chapter aims to review the recent development of MSA using meta-heuristics algorithms. In addition, two recent techniques are focused in more deep: the first is Fragmented protein sequence alignment using two-layer particle swarm optimization (FTLPSO). The second is Multiple sequence alignment using multi-objective based bacterial foraging optimization algorithm (MO-BFO).

A phase field modeling framework is developed to quantify structure evolution of protein fibrils in solution. The modeling framework employs a set of multi-physics constitutive relations to predict time dependent protein fibril structural evolution. The balance relations include chemical potential relations, microforces that govern local protein structure evolution, linear momentum and conservation of mass. Anisotropic formation of protein fibrils is controlled by protein monomer microforces and chemical fluxes to obtain long fibril growth from small seed particles. The theoretical model is implemented numerically using a nonlinear finite element phase field modeling approach which couples nonlinear mechanics with microscopic protein fibril structure evolution and chemical behavior. For comparisons to the model, the self-healing RADA16-I protein fibrils are characterized using transmission electron microscopy before and after ultrasonic radiation. Comparisons illustrate quantitative model predictions that govern spontaneous protein fibril self-healing that is predicted on the time scale of several hundred hours. The underlying physical mechanisms associated with self-assembly of the protein fibrils are discussed.

Here we focus on recent advances in understanding the deformation and fracture behavior of collagen, Nature’s most abundant protein material and the basis for many biological composites including bone, dentin or cornea. We show that it is due to the basis of the collagen structure that leads to its high strength and ability to sustain large deformation, as relevant to its physiological role in tissues such as bone and muscle. Experiment has shown that collagen isolated from different sources of tissues universally displays a design that consists of tropocollagen molecules with lengths of approximately 300 nanometers. Using a combination of theoretical analyses and multi-scale modeling, we have discovered that the characteristic structure and characteristic dimensions of the collagen nanostructure is the key to the ability to take advantage of the nanoscale properties of individual tropocollagen molecules at larger scales, leading to a tough material at the micro- and mesoscale. This is achieved by arranging tropocollagen molecules into a staggered assembly at a specific optimal molecular length scale. During bone formation, nanoscale mineral particles precipitate at highly specific locations in the collagen structure. These mineralized collagen fibrils are highly conserved, nanostructural primary building blocks of bone. By direct molecular simulation of the bone’s nanostructure, we show that it is due to the characteristic nanostructure of mineralized collagen fibrils that leads to its high strength and ability to sustain large deformation, as relevant to its physiological role, creating a strong and tough material. We present a thorough analysis of the molecular mechanisms of protein and mineral phases in deformation, and report discovery of a new fibrillar toughening mechanism that has major implications on the fracture mechanics of bone. Our studies of collagen and bone illustrate how hierarchical multi-scale modeling linking quantum chemistry with continuum fracture mechanics approaches can be used to develop predictive models of hierarchical protein materials. We conclude with a discussion of the significance of hierarchical multi-scale structures for the material properties and illustrate how these structures enable one to overcome some of the limitations of conventional materials design, combining disparate material properties such as strength and robustness.