Get5 Carboxyl-terminal Domain Is a Novel Dimerization Motif That Tethers an Extended Get4/Get5 Complex

Tail-anchored trans-membrane proteins are targeted to membranes post-translationally. The proteins Get4 and Get5 form an obligate complex that catalyzes the transfer of tail-anchored proteins destined to the endoplasmic reticulum from Sgt2 to the cytosolic targeting factor Get3. Get5 forms a homodimer mediated by its carboxyl domain. We show here that a conserved motif exists within the carboxyl domain. A high resolution crystal structure and solution NMR structures of this motif reveal a novel and stable helical dimerization domain. We additionally determined a solution NMR structure of a divergent fungal homolog, and comparison of these structures allows annotation of specific stabilizing interactions. Using solution x-ray scattering and the structures of all folded domains, we present a model of the full-length Get4/Get5 complex

A Structural Model of the Sgt2 Protein and Its Interactions with Chaperones and the Get4/Get5 Complex

The insertion of tail-anchored transmembrane (TA) proteins into the appropriate membrane is a post-translational event that requires stabilization of the transmembrane domain and targeting to the proper destination. Sgt2 is a heat-shock protein cognate (HSC) co-chaperone that preferentially binds endoplasmic reticulum-destined TA proteins and directs them to the GET pathway via Get4 and Get5. Here, we present the crystal structure from a fungal Sgt2 homolog of the tetratrico-repeat (TPR) domain and part of the linker that connects to the C-terminal domain. The linker extends into the two-carboxylate clamp of the TPR domain from a symmetry-related molecule mimicking the binding to HSCs. Based on this structure, we provide biochemical evidence that the Sgt2 TPR domain has the ability to directly bind multiple HSC family members. The structure allows us to propose features involved in this lower specificity relative to other TPR containing co-chaperones. We further show that a dimer of Sgt2 binds a single Get5 and use small angle x-ray scattering to characterize the domain arrangement of Sgt2 in solution. These results allow us to present a structural model of the Sgt2-Get4/Get5-HSC complex

Effects of seawater and deionized water at 0 to 80 deg C on the flexural properties of a glass/epoxy composite

The effect on the flexural properties of a glass/epoxy composite of immersion in deionized water or seawater at 0, 25, and 80 C for 451 hr was examined. The percent weight gain at 0 and 25 C was low (0.06 to 0.17 percent) and there was no significant change in the flexural properties for these environmental conditions. At 80 C there was a decrease in the flexural strength of 17 and 20 percent in seawater and deionized water, respectively. This is a comparison to control samples exposed to 80 C heat alone. These decreases were found to be nearly reversible once the samples were dried. Optical microscopy did not reveal cracking of the matrix. The flexural modulus was essentially unaffected by exposure to deionized water and seawater at 80 C

Model for eukaryotic tail-anchored protein binding based on the structure of Get3

The Get3 ATPase directs the delivery of tail-anchored (TA) proteins to the endoplasmic reticulum (ER). TA-proteins are characterized by having a single transmembrane helix (TM) at their extreme C terminus and include many essential proteins, such as SNAREs, apoptosis factors, and protein translocation components. These proteins cannot follow the SRP-dependent co-translational pathway that typifies most integral membrane proteins; instead, post-translationally, these proteins are recognized and bound by Get3 then delivered to the ER in the ATP dependent Get pathway. To elucidate a molecular mechanism for TA protein binding by Get3 we have determined three crystal structures in apo and ADP forms from Saccharomyces cerevisae (ScGet3-apo) and Aspergillus fumigatus (AfGet3-apo and AfGet3-ADP). Using structural information, we generated mutants to confirm important interfaces and essential residues. These results point to a model of how Get3 couples ATP hydrolysis to the binding and release of TA-proteins

Ultrastructure and complex polar architecture of the human pathogen Campylobacter jejuni

Campylobacter jejuni is one of the most successful food-borne human pathogens. Here we use electron cryotomography to explore the ultrastructure of C. jejuni cells in logarithmically growing cultures. This provides the first look at this pathogen in a near-native state at macromolecular resolution (~5 nm). We find a surprisingly complex polar architecture that includes ribosome exclusion zones, polyphosphate storage granules, extensive collar-shaped chemoreceptor arrays, and elaborate flagellar motors

Organization theory and military metaphor: time for a reappraisal?

A ‘conventional’ use of military metaphor would use it to convey attributes such as hierarchical organization, vertical communication and limited autonomy. This is often used in contrast to a looser form of organization based on the metaphor of the network. However, this article argues that military practice is more complex, with examples of considerable autonomy within the constraints of central direction. It is suggested that not only might this be a more useful metaphor for many contemporary organizations, but also that simplistic uses of military metaphor divert our attention away from the functions that management hierarchies play. The discussion is embedded within a critical realist account of metaphor, arguing for both its value and the need for its further development

Size-structured populations: immigration, (bi)stability and the net growth rate

We consider a class of physiologically structured population models, a first order nonlinear partial differential equation equipped with a nonlocal boundary condition, with a constant external inflow of individuals. We prove that the linearised system is governed by a quasicontraction semigroup. We also establish that linear stability of equilibrium solutions is governed by a generalized net reproduction function. In a special case of the model ingredients we discuss the nonlinear dynamics of the system when the spectral bound of the linearised operator equals zero, i.e. when linearisation does not decide stability. This allows us to demonstrate, through a concrete example, how immigration might be beneficial to the population. In particular, we show that from a nonlinearly unstable positive equilibrium a linearly stable and unstable pair of equilibria bifurcates. In fact, the linearised system exhibits bistability, for a certain range of values of the external inflow, induced potentially by All\'{e}e-effect.Comment: to appear in Journal of Applied Mathematics and Computin

Mechanism of assembly of a substrate-transfer complex during tail-anchored protein targeting

Tail-anchored (TA) proteins, defined as having a single transmembrane helix at their C terminus, are post-translationally targeted to the endoplasmic reticulum membrane by the guided entry of TA proteins (GET) pathway. In yeast, the handover of TA substrates is mediated by the heterotetrameric Get4/Get5 complex (Get4/5), which tethers the co-chaperone Sgt2 to the targeting factor, the Get3 ATPase. Binding of Get4/5 to Get3 is critical for efficient TA targeting; however, questions remain about the formation of the Get3·Get4/5 complex. Here we report crystal structures of a Get3·Get4/5 complex from Saccharomyces cerevisiae at 2.8 and 6.0 Å that reveal a novel interface between Get3 and Get4 dominated by electrostatic interactions. Kinetic and mutational analyses strongly suggest that these structures represent an on-pathway intermediate that rapidly assembles and then rearranges to the final Get3·Get4/5 complex. Furthermore, we provide evidence that the Get3·Get4/5 complex is dominated by a single Get4/5 heterotetramer bound to one monomer of a Get3 dimer, uncovering an intriguing asymmetry in the Get4/5 heterotetramer upon Get3 binding. Ultrafast diffusion-limited electrostatically driven Get3·Get4/5 association enables Get4/5 to rapidly sample and capture Get3 at different stages of the GET pathway

Atomic Structures of the 30S Subunit and Its Complexes with Ligands and Antibiotics

The two subunits that make up the ribosome have both distinct and cooperative functions. The 30S ribosomal subunit binds messenger RNA (mRNA) and is involved in the selection of cognate transfer RNA (tRNA) by monitoring codon–anticodon base-pairing during the decoding process. The 50S subunit catalyzes peptide-bond formation. Both subunits work in concert to move tRNAs and mRNAs relative to the ribosome in translocation, and both are the target of a large number of naturally occurring antibiotics. Thus, useful information about the mechanism of translation can be gleaned from structures of both individual subunits and the intact ribosome. In this paper, we describe our work on the determination of the atomic structure of the 30S ribosomal subunit and its complexes with RNA ligands, antibiotics, and initiation factor IF1. The results provide structural insights into how the ribosome recognizes cognate tRNA and discriminates against near-cognate tRNA. They also provide a structural basis for understanding the action of various antibiotics that target the 30S subunit

Communicating product user reviews and ratings in interfaces for e-commerce: a multimodal approach

This paper describes a comparative empirical evaluation study that uses multimodal presentations to communicate review messages in an e-commerce platform. Previous studies demonstrate the effective use of multimodality in different problem domains (e.g. e-learning). In this paper, multimodality and expressive avatars are used to communicate information related to product reviews messages. The data of the reviews was opportunistically collected from Facebook and Twitter. Two independent groups of users were used to evaluate two different presentations of reviews and ratings using as a basis an experimental e- commerce platform. The control group used a text-based with emojis presentation and the experimental group used a multimodal approach based on expressive avatars. Three parameters of usability were measured. These were efficiency, effectiveness, user satisfaction, and user preference. The result showed that the two approaches performed similarly. These findings provide a basis for further experiments in which text, emojis and expressive avatars can be combine to communicate a larger volume of reviews and ratings