scholarly journals The ins and outs of vesicular monoamine transporters

2018 ◽  
Vol 150 (5) ◽  
pp. 671-682 ◽  
Author(s):  
Dana Yaffe ◽  
Lucy R. Forrest ◽  
Shimon Schuldiner
Keyword(s):  
High Resolution ◽  
Conformational Equilibrium ◽  
Structural Data ◽  
Conformational Transitions ◽  
Vesicular Monoamine Transporter ◽  
Monoamine Transporters ◽  
Detailed Understanding ◽  
Transport Cycle ◽  
Vesicular Monoamine Transporters ◽  
Homology Models

The H+-coupled vesicular monoamine transporter (VMAT) is a transporter essential for life. VMAT mediates packaging of the monoamines serotonin, dopamine, norepinephrine, and histamine from the neuronal cytoplasm into presynaptic vesicles, which is a key step in the regulated release of neurotransmitters. However, a detailed understanding of the mechanism of VMAT function has been limited by the lack of availability of high-resolution structural data. In recent years, a series of studies guided by homology models has revealed significant insights into VMAT function, identifying residues that contribute to the binding site and to specific steps in the transport cycle. Moreover, to characterize the conformational transitions that occur upon binding of the substrate and coupling ion, we have taken advantage of the unique and powerful pharmacology of VMAT as well as of mutants that affect the conformational equilibrium of the protein and shift it toward defined conformations. This has allowed us to identify an important role for the proton gradient in driving a shift from lumen-facing to cytoplasm-facing conformations.

Expression and function of the rat vesicular monoamine transporter 2

2008 ◽  
Vol 294 (4) ◽  
pp. C1004-C1011 ◽  
Author(s):  
Yoav Adam ◽  
Robert H. Edwards ◽  
Shimon Schuldiner
Keyword(s):  
Structural Data ◽  
Monoamine Transporters ◽  
Substrate Molecule ◽  
Electrogenic Transport ◽  
Multidrug Transporters ◽  
The Central Nervous System ◽  
Vesicular Monoamine Transporters ◽  
Vesicular Transporters ◽  
High Level ◽  
And Function

The vesicular monoamine transporters (VMATs) are essential proteins, involved in the storage of monoamines in the central nervous system and in endocrine cells, in a process that involves exchange of 2H+with one substrate molecule. The VMATs interact with various native substrates and clinically relevant drugs and display the pharmacological profile of multidrug transporters. Vesicular transporters suffer from a lack of biochemical and structural data due to the difficulties in their expression. In this work we present the high-level expression of rat VMAT2 (rVMAT2) in a stable a human embryonic kidney cell line (HEK293), generated using the resistance to the neurotoxin 1-methyl-4-phenylpyridinium (MPP+) conferred by the protein. In addition, we describe novel procedures for the solubilization and purification of active protein, and its reconstitution into proteoliposomes. The partially purified protein in detergent binds the inhibitor tetrabenazine and, after reconstitution, displays high levels of ΔμH+-driven electrogenic transport of serotonin. The reconstituted purified rVMAT2 has wild-type affinity for serotonin, and its turnover rate is ∼0.4 substrate molecule/s.

Tubulin structure at intermediate resolution

1995 ◽  
Vol 53 ◽  
pp. 852-853
Author(s):  
K. H. Downing ◽  
S. G. Wolf ◽  
E. Nogales
Keyword(s):  
High Resolution ◽  
Structural Data ◽  
Therapeutic Drug ◽  
Zinc Ions ◽  
Two Dimensional ◽  
X Ray Diffraction ◽  
X Ray ◽  
Negative Stain ◽  
Functional Mechanisms ◽  
Tubulin Structure

Microtubules are involved in a host of critical cell activities, many of which involve transport of organelles through the cell. Different sets of microtubules appear to form during the cell cycle for different functions. Knowledge of the structure of tubulin will be necessary in order to understand the various functional mechanisms of microtubule assemble, disassembly, and interaction with other molecules, but tubulin has so far resisted crystallization for x-ray diffraction studies. Fortuitously, in the presence of zinc ions, tubulin also forms two-dimensional, crystalline sheets that are ideally suited for study by electron microscopy. We have refined procedures for forming the sheets and preparing them for EM, and have been able to obtain high-resolution structural data that sheds light on the formation and stabilization of microtubules, and even the interaction with a therapeutic drug.Tubulin sheets had been extensively studied in negative stain, demonstrating that the same protofilament structure was formed in the sheets and microtubules. For high resolution studies, we have found that the sheets embedded in either glucose or tannin diffract to around 3 Å.

HRTEM study of valleriite, a hydroxide-bearing copper sulfide

1990 ◽  
Vol 48 (4) ◽  
pp. 462-463
Author(s):  
S. Wang ◽  
P. R. Buseck
Keyword(s):  
Electron Microscope ◽  
High Resolution ◽  
Copper Sulfide ◽  
Carbonaceous Chondrite ◽  
Structural Data ◽  
Basic Structure ◽  
Long Period ◽  
Selected Area Electron Diffraction ◽  
Transmission Electron ◽  
Wavy Structure

Valleriite is an unusual mineral, consisting of intergrowths of sulfide layers (corresponding in structure to the mineral smythite - Fe9S11) and hydroxide layers (corresponding to brucite - Mg(OH2)). It has a composition of approximately 1.526[Mg.68Al.32(OH)2].[Fe1.07Cu.93S2] and consists of two interpenetrating lattices, each of which retains its individual structural and diffraction characteristics parallel to the layering. The valleriite structure is related to that of tochilinite, an unusual iron-rich mineral that is of considerable interest for the origin of certain carbonaceous chondrite meteorites and to those of franckeite and cylindrite, two minerals that are of interest because of their unique morphological and crystallographic properties, e.g., the distinctive curved form of cylindrite and the perfect mica-like cleavage with unusual striations and the long-period wavy structure of franckeite.Our selected-area electron diffraction (SAED) patterns and high-resolution transmission electron microscope (HRTEM) images of valleriite provide new structural data. A basic structure and a new superstructure have been observed.

scholarly journals Studies of Conformational Changes of Tubulin Induced by Interaction with Kinesin Using Atomistic Molecular Dynamics Simulations

2021 ◽  
Vol 22 (13) ◽  
pp. 6709
Author(s):  
Xiao-Xuan Shi ◽  
Peng-Ye Wang ◽  
Hong Chen ◽  
Ping Xie
Keyword(s):  
Molecular Dynamics ◽  
High Resolution ◽  
Binding Energy ◽  
Molecular Dynamics Simulations ◽  
Conformational Changes ◽  
Weak Interactions ◽  
Molecular Motor ◽  
Structural Data ◽  
Calculated Binding Energy ◽  
Dynamics Simulations

The transition between strong and weak interactions of the kinesin head with the microtubule, which is regulated by the change of the nucleotide state of the head, is indispensable for the processive motion of the kinesin molecular motor on the microtubule. Here, using all-atom molecular dynamics simulations, the interactions between the kinesin head and tubulin are studied on the basis of the available high-resolution structural data. We found that the strong interaction can induce rapid large conformational changes of the tubulin, whereas the weak interaction cannot. Furthermore, we found that the large conformational changes of the tubulin have a significant effect on the interaction of the tubulin with the head in the weak-microtubule-binding ADP state. The calculated binding energy of the ADP-bound head to the tubulin with the large conformational changes is only about half that of the tubulin without the conformational changes.

scholarly journals Genetic or Toxicant-Induced Disruption of Vesicular Monoamine Storage and Global Metabolic Profiling in Caenorhabditis elegans

2021 ◽  
Author(s):  
Joshua M Bradner ◽  
Vrinda Kalia ◽  
Fion K Lau ◽  
Monica Sharma ◽  
Meghan L Bucher ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Caenorhabditis Elegans ◽  
High Resolution ◽  
High Resolution Mass Spectrometry ◽  
Dopamine Neurons ◽  
Vesicular Monoamine Transporter ◽  
Monoamine Transporter ◽  
C Elegans ◽  
High Resolution Mass ◽  
Resolution Mass

Abstract The proper storage and release of monoamines contributes to a wide range of neuronal activity. Here, we examine the effects of altered vesicular monoamine transport in the nematode Caenorhabditis elegans. The gene cat-1 is responsible for the encoding of the vesicular monoamine transporter (VMAT) in C. elegans and is analogous to the mammalian vesicular monoamine transporter 2 (VMAT2). Our laboratory has previously shown that reduced VMAT2 activity confers vulnerability on catecholamine neurons in mice. The purpose of this article was to determine whether this function is conserved and to determine the impact of reduced VMAT activity in C. elegans. Here we show that deletion of cat-1/VMAT increases sensitivity to the neurotoxicant 1-methyl-4-phenylpyridinium (MPP+) as measured by enhanced degeneration of dopamine neurons. Reduced cat-1/VMAT also induces changes in dopamine-mediated behaviors. High-resolution mass spectrometry-based metabolomics in the whole organism reveals changes in amino acid metabolism, including tyrosine metabolism in the cat-1/VMAT mutants. Treatment with MPP+ disrupted tryptophan metabolism. Both conditions altered glycerophospholipid metabolism, suggesting a convergent pathway of neuronal dysfunction. Our results demonstrate the evolutionarily conserved nature of monoamine function in C. elegans and further suggest that high-resolution mass spectrometry-based metabolomics can be used in this model to study environmental and genetic contributors to complex human disease.

High-resolution Measurements of the Galactic Radio Spectrum

1969 ◽  
Vol 1 (5) ◽  
pp. 210-211 ◽  
Author(s):  
T. L. Landecker ◽  
R. Wielebinski
Keyword(s):  
High Resolution ◽  
Radio Emission ◽  
High Latitude ◽  
Galactic Plane ◽  
Background Radiation ◽  
Radio Spectrum ◽  
Synchrotron Emission ◽  
Detailed Structure ◽  
Detailed Understanding ◽  
Galactic Radio

High-resolution surveys of the galactic radio emission show detailed structure both near the galactic plane and at high galactic latitudes. Some of this structure is describable as ‘spurs’ while the rest comprises isolated features. We describe here measurements of the spectral index of the background radiation with a resolution of 3°.5. These measurements are aimed at a detailed understanding of the high-latitude synchrotron emission.

The mechanism of the releasing action of amphetamine. Uptake, superfusion, and electrophysiological studies on transporter-transfected cells

2000 ◽  
Vol 72 (6) ◽  
pp. 1045-1050 ◽  
Author(s):  
C. Pifl ◽  
H. H Sitte ◽  
H. Reither ◽  
E. A. Singer
Keyword(s):  
Patch Clamp ◽  
Molecular Targets ◽  
Monoamine Transporters ◽  
Transfected Cells ◽  
Whole Cell Patch Clamp ◽  
Sodium Dependent ◽  
Electrophysiological Studies ◽  
Inward Currents ◽  
Vesicular Monoamine Transporters ◽  
The Brain

Amphetamine analogues are able to induce signs of neurotoxicity in the brain. In order to understand this type of neurotoxicity, the interaction of amphetamine with its molecular targets must be elucidated. These molecular targets are plasmalemmal and vesicular monoamine transporters. We investigated the interaction of amphetamine with these transporters in cells transfected with the respective cDNA. Superfusion and whole-cell, patch-clamp experiments were performed, and the toxicity of substrates of the transporters was studied. Amphetamine was taken up by dopamine transporter-expressing cells in a sodium-dependent and cocaine-blockable manner. Furthermore, it elicited inward currents in these cells concentration-dependently. Correlation of uptake, release, and patch-clamp experiments suggest that ion fluxes induced by substrate-gating on transporters may significantly contribute to the releasing action of amphetamine and of other transporter substrates. Dopamine accumulation into serotoninergic terminals depleted of serotonin by 3,4-methylenedioxymethamphetamine was discussed as a mechanism of Ecstasy-toxicity. This is in agreement with a toxic effect of intracellular dopamine which could be demonstrated on our transporter-overexpressing cells. These results, apart from their relevance for the toxicity by amphetamine analogues, may also have bearings on the mechanisms in neurodegenerative diseases affecting monoamine transmitters.

scholarly journals The High Resolution Crystal Structure of the Human Tumor Suppressor Maspin Reveals a Novel Conformational Switch in the G-helix

2005 ◽  
Vol 280 (23) ◽  
pp. 22356-22364 ◽  
Author(s):  
Ruby H. P. Law ◽  
James A. Irving ◽  
Ashley M. Buckle ◽  
Katya Ruzyla ◽  
Marguerite Buzza ◽  
...  
Keyword(s):  
Crystal Structure ◽  
High Resolution ◽  
Tumor Suppressor ◽  
Physiological Role ◽  
Human Tumor ◽  
Salt Bridge ◽  
Structural Data ◽  
Crystal Forms ◽  
Reactive Center ◽  
Α Helix

Maspin is a serpin that acts as a tumor suppressor in a range of human cancers, including tumors of the breast and lung. Maspin is crucial for development, because homozygous loss of the gene is lethal; however, the precise physiological role of the molecule is unclear. To gain insight into the function of human maspin, we have determined its crystal structure in two similar, but non-isomorphous crystal forms, to 2.1- and 2.8-Å resolution, respectively. The structure reveals that maspin adopts the native serpin fold in which the reactive center loop is expelled fully from the A β-sheet, makes minimal contacts with the core of the molecule, and exhibits a high degree of flexibility. A buried salt bridge unique to maspin orthologues causes an unusual bulge in the region around the D and E α-helices, an area of the molecule demonstrated in other serpins to be important for cofactor recognition. Strikingly, the structural data reveal that maspin is able to undergo conformational change in and around the G α-helix, switching between an open and a closed form. This change dictates the electrostatic character of a putative cofactor binding surface and highlights this region as a likely determinant of maspin function. The high resolution crystal structure of maspin provides a detailed molecular framework to elucidate the mechanism of function of this important tumor suppressor.

Sign in / Sign up

Export Citation Format

Share Document