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

Microstructural and mechanical behavior of Al 6061/SiC-Al2O3 composites processed through friction stir processing

Metal Matrix Composite (MMC) reinforced in friction stir processing (FSP) has increased insights that can affectively attain the desired mechanical properties for the manufactured samples. The favorable conditions of carbides are considered for reinforcing the SiC particles into the Aluminum 6061. The methodology of fabricating Aluminum 6061 comprises of three materials, Al 6061-SiC-Al2O3. The experimental evaluation of the composite Aluminum 6061-SiC-Al2O3 includes the influence of process parameters on microhardness, tensile strength, and microstructure. As a result of the reinforcement of nanoparticles processed in FSP, the properties of composite material increased satisfactorily. The sample S3 observed to be having a maximum tensile strength of 185 MPa. The larger, the better condition is adopted to analyze the tensile strength of the fabricated samples. The optimum condition for maximum tensile strength was found at 900 RPM, 15 mm/min, and composition 3. The hardness profiles at different zones of friction stir processing (FSP), viz., Heat Affected Zone (HAZ), Thermo Mechanical Affected Zone (TMAZ), Nugget Zone (NZ) were examined. The characterization techniques deployed were optical microscope (OM), and scanning electron microscope (SEM) studies for microstructural behavior. The result shows that the reinforcements were tightly embedded into the base material surface. The spherical grains are formed in the reinforcement region

Mechanisms of Regulation and Fidelity in Tail-Anchored Membrane Protein Targeting

Accurate protein localization is crucial to generate and to maintain cellular organization. Achieving accuracy is challenging, as the molecular signals that dictate a protein’s destination are often promiscuous. The localization of tail-anchored (TA) proteins, whose transmembrane domain resides at its extreme C-terminus, presents major challenges to protein targeting machineries. This dissertation explores how TA capture and release are spatially and temporally regulated in the Guided Entry of Tail Anchored proteins (GET) pathway and how endoplasmic reticulum (ER) destined TAs are targeted with high fidelity. A quantitative framework of the Get3 ATPase cycle reveals that ATP and GET pathway effector proteins specifically induce multiple conformational changes in Get3, which culminate in the ATPase activation that drives unidirectional targeting in the pathway. The Get4/5 TA loading complex locks Get3 in the ATP-bound state that is primed for TA protein capture, whereas the TA substrate induces tetramerization of Get3 and activates its ATPase reaction. Additional analyses define multiple physicochemical features that distinguish TA proteins destined to different organelles. The GET pathway selects for these features at distinct stages using mechanisms such as differential binding, induced fit, and kinetic proofreading after ATP hydrolysis by Get3. These results reveal new roles for the cochaperone Sgt2 in providing key selection filters, and provide a biological logic for the complex cascade of substrate relay events during post-translational membrane protein targeting.</p

Precise Timing of ATPase Activation Drives Targeting of Tail-Anchored Proteins

Hundreds of proteins are anchored on cellular membranes by a transmembrane domain (TM) at their extreme C-terminus. These ‘tail-anchored' (TA) proteins perform numerous essential cellular functions, yet their unique topology poses fundamental challenges to their proper localization. In eukaryotic cells, the highly conserved ATPase Get3 coordinates the efficient capture and delivery of TA proteins to the endoplasmic reticulum (ER). How Get3 uses its ATPase cycles to drive this fundamental process remains elusive. Here we define the Get3 ATPase cycle and show that ATP specifically induces conformational changes in Get3 that culminate in its ATPase activation through tetramerization. This activation is inhibited by the TA protein loading complex Get4/5 and is activated by the TA substrate, ensuring the precise timing of ATP hydrolysis during targeting. Our results provide an explicit model for how Get3 harnesses the energy from ATP to spatially and temporally coordinate the membrane localization of TA proteins

In vitro availability of essential amino acids, and the protein efficiency ration of cooked cicer arietinum

The legume Cicer arietinum (chick-pea) is a commonly used source of dietary protein in India because of its high biological value and non-toxic nature. The objective of this investigation was to study the effects of cooking on the availability of four essential amino acids in chick-peas and on the utilization of its protein for tissue synthesis. Acid hydrolysis for varying periods of time was used to determine the optimum level of each amino acid present in raw, roasted, boiled, and steam-cooked chick-peas. Enzymatic hydrolysis with pepsin and pancreatin for varying periods of time was used to study the quantitative release of the amino acids in vitro. The lysine, threonine, valine, and leucines present in the acid and enzyme hydrolysates were identified by 1-dimensional polychromatic paper chromatography using n-butanol, acetic acid, and water as the solvent for resolution. Quantitative estimations were based on the elution and colorimetric readings of the chromatograms. A growth experiment with weanling rats was employed to evaluate the overall quality of the protein. Using a randomized block design, five groups of 3 week old male rats of the Holtzman strain were fed a basal diet supplying 10% protein through casein, and experimental diets supplying 10% protein through raw, roasted, boiled, and steam-cooked chick-peas for a 4 week period

Differential gradients of interaction affinities drive efficient targeting and recycling in the GET pathway

Efficient and accurate localization of membrane proteins requires a complex cascade of interactions between protein machineries. This requirement is exemplified in the guided entry of tail-anchored (TA) protein (GET) pathway, where the central targeting factor Get3 must sequentially interact with three distinct binding partners to ensure the delivery of TA proteins to the endoplasmic reticulum (ER) membrane. To understand the molecular principles that provide the vectorial driving force of these interactions, we developed quantitative fluorescence assays to monitor Get3–effector interactions at each stage of targeting. We show that nucleotide and substrate generate differential gradients of interaction energies that drive the ordered interaction of Get3 with successive effectors. These data also provide more molecular details on how the targeting complex is captured and disassembled by the ER receptor and reveal a previously unidentified role for Get4/5 in recycling Get3 from the ER membrane at the end of the targeting reaction. These results provide general insights into how complex protein interaction cascades are coupled to energy inputs in biological systems

Open Neighborhood Coloring of Prisms

For a simple, connected, undirected graph G(V, E) an open neighborhood coloring of the graph G is a mapping f : V (G) --&gt; Z+ such that for each w in V(G), and for all u, v in N(w), f(u) is different from f(v). The maximum value of f(w), for all w in V (G) is called the span of the open neighborhood coloring f. The minimum value of span of f over all open neighborhood colorings f is called open neighborhood chromatic number of G, denoted by Xonc(G). In this paper we determine the open neighborhood chromatic number of prisms

Multiple selection filters ensure accurate tail-anchored membrane protein targeting

Accurate protein localization is crucial to generate and maintain organization in all cells. Achieving accuracy is challenging, as the molecular signals that dictate a protein's cellular destination are often promiscuous. A salient example is the targeting of an essential class of tail-anchored (TA) proteins, whose sole defining feature is a transmembrane domain near their C-terminus. Here we show that the Guided Entry of Tail-anchored protein (GET) pathway selects TA proteins destined to the endoplasmic reticulum (ER) utilizing distinct molecular steps, including differential binding by the co-chaperone Sgt2 and kinetic proofreading after ATP hydrolysis by the targeting factor Get3. Further, the different steps select for distinct physicochemical features of the TA substrate. The use of multiple selection filters may be general to protein biogenesis pathways that must distinguish correct and incorrect substrates based on minor differences

Histo-pathological findings in kidneys with polar artery: a demystifying endeavor

Background: The kidneys are supplied by renal arteries which enter the hilum and branch progressively from the pyramids to the cortex into lobar, interlobar, arcuate, interlobar, interlobular, and finally terminate as afferent arterioles that enter the glomeruli. Thus there is a normal pattern of blood flow towards the cortex from the pyramids when the artery enters at the hilum. The aim of the study was to explore the glomeruli pattern in kidneys with polar arteries.Methods: The study was conducted in the department of anatomy at a tertiary care referral institute. Twenty-two kidneys with polar arteries were obtained by conventional method of dissection from cadavers for the study. Sections were taken from the upper pole and lower pole in each kidney. The sections were taken perpendicular and close to the polar arteries. The sections were subjected to routine histological processing and staining as per the standard operating procedure. Histological findings were observed and documented.Results: The study found that the kidneys were histologically different with polar arteries as compared to normal kidneys. The number of glomeruli per high power field was higher nearer to the polar arteries. The glomeruli were viable and not sclerosed. An agglomeration of arterioles were present close to the polar arteries but they were distinct and did not seem to be associated with glomeruli.    Conclusions: The study found an agglomeration of arterioles, increased number of viable glomeruli and cystic changes associated with kidneys having polar artery. Thus this warrants a detailed study with special stains for research to elucidate the mechanisms of the circulation in polar arteries and correlation of the same findings with clinical conditions such as hypertension or any other diseases of the kidney

Concurrent homozygous sickle‐cell disease and severe haemophilia A: Thromboelastography profiles

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/148348/1/hae13692_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/148348/2/hae13692.pd