Structure of the integrin binding fragment from fibrillin-1 gives new insights into microfibril organization

Human fibrillin-1, the major structural protein of extracellular matrix (ECM) 10-12 nm microfibrils, is dominated by 43 calcium binding epidermal growth factor-like (cbEGF) and 7 transforming growth factor beta binding protein-like (TB) domains. Crystal structures reveal the integrin binding cbEGF22-TB4-cbEGF23 fragment of human fibrillin-1 to be a Ca(2+)-rigidified tetragonal pyramid. We suggest that other cbEGF-TB pairs within the fibrillins may adopt a similar orientation to cbEGF22-TB4. In addition, we have located a flexible RGD integrin binding loop within TB4. Modeling, cell attachment and spreading assays, immunocytochemistry, and surface plasmon resonance indicate that cbEGF22 bound to TB4 is a requirement for integrin activation and provide insight into the molecular basis of the fibrillin-1 interaction with alphaVbeta3. In light of our data, we propose a novel model for the assembly of the fibrillin microfibril and a mechanism to explain its extensibility.     

Glycine receptors: recent insights into their structural organization and functional diversity

Strychnine-sensitive glycine receptors (GlyRs) are known to mediate synaptic inhibition in spinal cord, brainstem and other regions of the CNS. During the past 5 years, considerable progress has been made in delineating structural determinants of ligand binding and channel activation in recombinant GlyRs. Furthermore, immunohistochemical and gene inactivation studies have disclosed distinct distributions and functions of differentially expressed GlyR subtypes in retina, hippocampus and the dorsal horn of the spinal cord. Accordingly, GlyRs regulate not only the excitability of motor and sensory neurones, but are also essential for the processing of photoreceptor signals, neuronal development and inflammatory pain sensitization. Hence, these receptors constitute promising targets for the development of clinically useful compounds.     

Lactoperoxidase: structural insights into the function,ligand binding and inhibition

Lactoperoxidase (LPO) is a member of a large group of mammalian heme peroxidases that include myeloperoxidase (MPO), eosinophil peroxidase (EPO) and thyroid peroxidase (TPO). The LPO is found in exocrine secretions including milk. It is responsible for the inactivation of a wide range of micro-organisms and hence, is an important component of defense mechanism in the body. With the help of hydrogen peroxide, it catalyzes the oxidation of halides, pseudohalides and organic aromatic molecules. Historically, LPO was isolated in 1943, nearly seventy years ago but its three-dimensional crystal structure has been elucidated only recently. This review provides various details of this protein from its discovery to understanding its structure, function and applications. In order to highlight species dependent variations in the structure and function of LPO, a detailed comparison of sequence, structure and function of LPO from various species have been made. The structural basis of ligand binding and distinctions in the modes of binding of substrates and inhibitors have been analyzed extensively.     

Cleave to leave: structural insights into the dynamic organization of the nuclear pore complex

A detailed understanding of the fine structure of the nuclear pore complex has remained elusive. Now, studies on a small protein domain have shed light on the dynamic organization of this massive assembly.     

Dissecting the mammalian genome--new insights into chromosomal evolution

A recent study aligning genomic data from eight mammalian species has provided new and detailed information on the architecture of chromosomes that are thought to comprise the karyotype of the boreoeutherian ancestor. The analyses suggest that evolutionary breakpoints are clustering in "hotspots", that these regions are enriched for centromeres and that the more commonly occurring human cancer-associated breakpoints tend to co-localize with evolutionary breakpoints.     

New insights into the structural organization of eukaryotic and prokaryotic cytoskeletons using cryo-electron tomography

Cryo-electron tomography (cryo-ET) is an emerging imaging technology that combines the potential of three-dimensional (3-D) imaging at molecular resolution (<5 nm) with a close-to-life preservation of the specimen. In conjunction with pattern recognition techniques, it enables us to map the molecular landscape inside cells. The application of cryo-ET to intact cells provides novel insights into the structure and the spatial organization of the cytoskeleton in prokaryotic and eukaryotic cells.     

Current insights into chromatin structure organization

This review summarizes current insights into organization of chromatin structure at different levels of DNA compaction. Analysis of available experimental data allowed concluding that only nucleosomal level of structural organization was sufficiently investigated, whereas structure of a 30-nm chromatin fiber remains an open issue. The data on the chromatin structure obtained at the level of the nucleus speak in favor of a biphasic fractal organization of chromatin.     

Evidence for the intramolecular pleating model of fibrillin microfibril organisation from single particle image analysis

Fibrillin microfibrils endow mammalian connective tissues with elasticity and are fundamental for the deposition of elastin. The microfibrils are 57nm periodic supramolecular protein polymers with a mass of 2.4MDa per repeat. The detailed structure and organisation of most matrix assemblies is poorly understood due to their large size and complexity and it has proved a major challenge to define their structural organisation. Therefore, we have used low dose electron microscopy and single particle image analysis to study the structure of fibrillin microfibrils. Three novel features were detected: a globular feature that bridges the "arm" region, a double band of density crossing the microfibril and stain penetrating holes present in the interbead region, possibly produced by the removal of microfibril associated proteins in the purification procedure. Fine filaments of approximately 2.4nm diameter are resolved in the interbead region, which correspond to the reported diameter of the fibrillin molecule. Comparison of the stain exclusion pattern of microfibrils with the theoretical stain exclusion pattern of fibrillin packing models indicates that the intramolecular pleating model, where each fibrillin molecule is pleated within one microfibril period allowing extensibility by unpleating, has the best fit to the data.     

Apolipoprotein E structure: insights into function

Human apolipoprotein E (apoE) is a member of the family of soluble apolipoproteins. Through its interaction with members of the low-density lipoprotein receptor family, apoE has a key role in lipid transport both in the plasma and in the central nervous system. Its three common structural isoforms differentially affect the risk of developing atherosclerosis and neurodegenerative disorders, including Alzheimer's disease. Because the function of apoE is dictated by its structure, understanding the structural properties of apoE and its isoforms is required both to determine its role in disease and for the development of therapeutic strategies.     

Emerging structural insights into C-type glycosyltransferases

Glycosyltransferases of the C superfamily (GT-Cs) are enzymes found in all domains of life. They catalyse the stepwise synthesis of oligosaccharides or the transfer of assembled glycans from lipid-linked donor substrates to acceptor proteins. The processes mediated by GT-Cs are required for C-, N- and O-linked glycosylation, all of which are essential post-translational modifications in higher-order eukaryotes. Until recently, GT-Cs were thought to share a conserved structural module of 7 transmembrane helices; however, recently determined GT-C structures revealed novel folds. Here we analyse the growing diversity of GT-C folds and discuss the emergence of two subclasses, termed GT-C_A and GT-C_B. Further substrate-bound structures are needed to facilitate a molecular understanding of glycan recognition and catalysis in these two subclasses.     

Protein networking: insights into global functional organization of proteomes

The formulation of network models from global protein studies is essential to understand the functioning of organisms. Network models of the proteome enable the application of Complex Network Analysis, a quantitative framework to investigate large complex networks using techniques from graph theory, statistical physics, dynamical systems and other fields. This approach has provided many insights into the functional organization of the proteome so far and will likely continue to do so. Currently, several network concepts have emerged in the field of proteomics. It is important to highlight the differences between these concepts, since different representations allow different insights into functional organization. One such concept is the protein interaction network, which contains proteins as nodes and undirected edges representing the occurrence of binding in large-scale protein-protein interaction studies. A second concept is the protein-signaling network, in which the nodes correspond to levels of post-translationally modified forms of proteins and directed edges to causal effects through post-translational modification, such as phosphorylation. Several other network concepts were introduced for proteomics. Although all formulated as networks, the concepts represent widely different physical systems. Therefore caution should be taken when applying relevant topological analysis. We review recent literature formulating and analyzing such networks.     

The spindle checkpoint: structural insights into dynamic signalling

Chromosome segregation is a complex and astonishingly accurate process whose inner working is beginning to be understood at the molecular level. The spindle checkpoint plays a key role in ensuring the fidelity of this process. It monitors the interactions between chromosomes and microtubules, and delays mitotic progression to allow extra time to correct defects. Here, we review and integrate findings on the dynamics of checkpoint proteins at kinetochores with structural information about signalling complexes.     

Outer membrane translocons: structural insights into channel formation

Gram-negative bacteria need to maintain the integrity of their outer membrane while also regulating the secretion of toxins and other macromolecules. A variety of dedicated outer membrane proteins (OMPs) facilitate this process. Recent structural work has shown that some of these proteins adopt classical β-barrel transmembrane structures and rely on structural changes within the barrel lumen to allow passage of substrate proteins. Other secretion systems have OMP components which use transmembrane α-helices and appear to function in a different way. Here we review a selection of recent structural studies which have major ramifications for our understanding of the passage of macromolecules across the outer membrane.     

Biogenesis and function of fibrillin assemblies

Fibrillin-1 and fibrillin-2 are large cysteine-rich glycoproteins that serve two key physiological functions: as supporting structures that impart tissue integrity and as regulators of signaling events that instruct cell performance. The structural role of fibrillins is exerted through the temporal and hierarchical assembly of microfibrils and elastic fibers, whereas the instructive role reflects the ability of fibrillins to sequester transforming growth factor β (TGFβ) and bone morphogenetic protein (BMP) complexes in the extracellular matrix. Characterization of fibrillin mutations in human patients and in genetically engineered mice has demonstrated that perturbation of either function manifests in disease. More generally, these studies have indicated that fibrillins are integral components of a broader biological network of extracellular, cell surface, and signaling molecules that orchestrate morphogenetic and homeostatic programs in multiple organ systems. They have also suggested that the relative composition of fibrillin-rich microfibrils imparts contextual specificity to TGFβ and BMP signaling by concentrating the ligands locally so as to regulate cell differentiation within a spatial context during organ formation (positive regulation) and by restricting their bioavailability so as to modulate cell performance in a timely fashion during tissue remodeling/repair (negative regulation). Correlative evidence suggests functional coupling of the cell-directed assembly of microfibrils and targeting of TGFβ and BMP complexes to fibrillins. Hence, the emerging view is that fibrillin-rich microfibrils are molecular integrators of structural and instructive signals, with TGFβ and BMPs as the nodal points that convert extracellular inputs into discrete and context-dependent cellular responses.     

Molecular insights into the ligand-controlled organization of the SAM-I riboswitch

S-adenosylmethionine (SAM) riboswitches are widespread in bacteria, and up to five different SAM riboswitch families have been reported, highlighting the relevance of SAM regulation. On the basis of crystallographic and biochemical data, it has been postulated, but never demonstrated, that ligand recognition by SAM riboswitches involves key conformational changes in the RNA architecture. We show here that the aptamer follows a two-step hierarchical folding selectively induced by metal ions and ligand binding, each of them leading to the formation of one of the two helical stacks observed in the crystal structure. Moreover, we find that the anti-antiterminator P1 stem is rotated along its helical axis upon ligand binding, a mechanistic feature that could be common to other riboswitches. We also show that the nonconserved P4 helical domain is used as an auxiliary element to enhance the ligand-binding affinity. This work provides the first comprehensive characterization, to our knowledge, of a ligand-controlled riboswitch folding pathway.