16 research outputs found
High‐Resolution NMR Determination of the Dynamic Structure of Membrane Proteins
No full textN-15 spin-relaxation rates are demonstrated to provide critical information about the long-range structure and internal motions of membrane proteins. Combined with an improved calculation method, the relaxation-rate-derived structure of the 283-residue human voltage-dependent anion channel revealed an anisotropically shaped barrel with a rigidly attached N-terminal helix. Our study thus establishes an NMR spectroscopic approach to determine the structure and dynamics of mammalian membrane proteins at high accuracy and resolution
Letter from Tsuneo Iwata to Mr. Harry L. Villinger, April 14, 1942
No full textLetter of gratitude from Tsuneo Iwata president of the Turlock Social Club to Mr. Harry L. Villinger, in response to the mass removal in California.The Nisaburo Aibara Collection features materials from the Turlock Social Club, a local Japanese-American community group active between 1939 and 1970. It contains documents regarding the Stockton, Turlock and Merced Assembly Centers and Japanese American Citizens League chapters. The Collection also features correspondences with reactions, responses, and preparations for the forced evacuation. Additionally, the Collection has records on the Central California Cantaloupe Company, Turlock Farm Corporation, Turlock Japanese Society, and family records and funeral service programs of Japanese-American residents of Turlock
Nucleotide interactions of the human voltage-dependent anion channel.
No full textBackground: Human VDAC1 mediates and controls the transport of metabolites across the outer mitochondrial membrane. Results: The N-terminal helix of hVDAC1 is involved in binding to charged forms of ATP, UTP, and GTP with an important contribution from lysine 20. Conclusion: Weak binding of ATP confers specificity for ATP transport. Significance: ATP interaction mapped at residue resolution supports metabolite selectivity of VDAC. The voltage-dependent anion channel (VDAC) mediates and gates the flux of metabolites and ions across the outer mitochondrial membrane and is a key player in cellular metabolism and apoptosis. Here we characterized the binding of nucleotides to human VDAC1 (hVDAC1) on a single-residue level using NMR spectroscopy and site-directed mutagenesis. We find that hVDAC1 possesses one major binding region for ATP, UTP, and GTP that partially overlaps with a previously determined NADH binding site. This nucleotide binding region is formed by the N-terminal -helix, the linker connecting the helix to the first -strand and adjacent barrel residues. hVDAC1 preferentially binds the charged forms of ATP, providing support for a mechanism of metabolite transport in which direct binding to the charged form exerts selectivity while at the same time permeation of the Mg2+-complexed ATP form is possible
Molecular Plasticity of the Human Voltage-Dependent Anion Channel Embedded Into a Membrane
No full textThe voltage-dependent anion channel (VDAC) regulates the flux of metabolites and ions across the outer mitochondrial membrane. Regulation of ion flow involves conformational transitions in VDAC, but the nature of these changes has not been resolved to date. By combining single-molecule force spectroscopy with nuclear magnetic resonance spectroscopy we show that the β barrel of human VDAC embedded into a membrane is highly flexible. Its mechanical flexibility exceeds by up to one order of magnitude that determined for β strands of other membrane proteins and is largest in the N-terminal part of the β barrel. Interaction with Ca2+, a key regulator of metabolism and apoptosis, considerably decreases the barrel's conformational variability and kinetic free energy in the membrane. The combined data suggest that physiological VDAC function depends on the molecular plasticity of its channel
Structure of the human voltage-dependent anion channel.
No full textThe voltage-dependent anion channel (VDAC), also known as mitochondrial porin, is the most abundant protein in the mitochondrial outer membrane (MOM). VDAC is the channel known to guide the metabolic flux across the MOM and plays a key role in mitochondrially induced apoptosis. Here, we present the 3D structure of human VDAC1, which was solved conjointly by NMR spectroscopy and x-ray crystallography. Human VDAC1 (hVDAC1) adopts a amp;946;-barrel architecture composed of 19 amp;946;-strands with an amp;945;-helix located horizontally midway within the pore. Bioinformatic analysis indicates that this channel architecture is common to all VDAC proteins and is adopted by the general import pore TOM40 of mammals, which is also located in the MOM
Functional dynamics in the voltage-dependent anion channel
No full textThe voltage-dependent anion channel (VDAC), located in the outer mitochondrial membrane, acts as a gatekeeper for the entry and exit of mitochondrial metabolites. Here we reveal functional dynamics of isoform one of VDAC (VDAC1) by a combination of solution NMR spectroscopy, Gaussian network model analysis, and molecular dynamics simulation. Micro- to millisecond dynamics are significantly increased for the N-terminal six ß-strands of VDAC1 in micellar solution, in agreement with increased B-factors observed in the same region in the bicellar crystal structure of VDAC1. Molecular dynamics simulations reveal that a charge on the membrane-facing glutamic acid 73 (E73) accounts for the elevation of N-terminal protein dynamics as well as a thinning of the nearby membrane. Mutation or chemical modification of E73 strongly reduces the micro- to millisecond dynamics in solution. Because E73 is necessary for hexokinase-I-induced VDAC channel closure and inhibition of apoptosis, our results imply that microto millisecond dynamics in the N-terminal part of the barrel are essential for VDAC interaction and gating
Untersuchung von Dynamik und Interaktionen des spannungsabhängigen Anionenkanals 1 mithilfe von NMR-Spektroskopie
No full textDer spannungsabhängige Anionenkanal (engl. “voltage-dependent anion channel”, VDAC), das häufigste Protein in der äußeren Mitochondrienmembran, dient als wichtiger Kontrollpunkt für den Ein- und Austritt von mitochondrialen Metaboliten und ist in mitochondriale Apoptose involviert. Die kürzliche Bestimmung der hochaufgelösten Struktur durch drei unabhängige Gruppen weist ein 19-strängiges β-Fass mit einer unterschiedlich arrangierten N-terminalen α-Helix in dessen Pore auf. In dieser Arbeit wird die Resonanzzuordnung der Isoform eins des humanen VDAC (VDAC1) in Lösung erweitert. Des Weiteren werden Nachweise für eine geknickte α-helikale Struktur des N-Terminus erbracht, die mit der Kristallstruktur von VDAC1 kompatibel ist, obwohl andere Strukturen nicht ausgeschlossen werden können. Zudem zeigt diese Studie funktionelle Dynamik in VDAC1 mithilfe einer Kombination aus Lösungs-NMR-Spektroskopie, Analyse Gauss’scher Netzwerkmodelle (GNM) und Molekulardynamik-(MD)-Simulationen. Niedrige Signalintensitäten deuten auf das Vorhandensein von Konformationsaustausch im zweiten Teil der N-terminalen α-Helix und dem Linker hin, der die α-Helix mit dem ersten β-Strang verbindet. Zusätzlich beeinflusst die Mutation von Arginin 15 im zweiten α-helikalen Teil die Stabilität der α-Helix und des gesamten β-Fasses in komplexer Weise. In Mizellen ist die Mikro- bis Millisekunden-Dynamik in der N-terminalen α-Helix, dem Linker und den N-terminalen sechs β-Strängen von VDAC1 signifikant erhöht. Außerdem sind die Wasserstoffbrücken der N-terminalen drei β-Stränge instabil. Übereinstimmend zeigen die N-terminalen Stränge erhöhte B-Faktoren in der Kristallstruktur von VDAC1 und durch GNM-Analyse vorhergesagte intrinsische Instabilität. Mutation oder chemische Modifizierung der in die Membran weisenden Glutaminsäure E73 reduzieren die in Lösung auftretende Mikro- bis Millisekunden-Dynamik nachhaltig. MD-Simulationen zeigen, dass eine Ladung an der Seitenkette von E73 für die Erhöhung der N-terminalen Proteindynamik sowie für eine Reduktion der Membrandicke in der Umgebung von E73 verantwortlich ist. Da E73 für die durch Hexokinase induzierte Kanalschließung und Inhibierung von Apoptose notwendig ist, zeigen diese Ergebnisse, dass die Mikro- bis Millisekunden-Dynamik im N-terminalen β-Fassbereich essentiell für Interaktionen und Kanalschließung von VDAC1 ist. Außerdem weisen die Daten auf einen Zusammenhang der Helix-Dynamik mit diesen Prozessen hin. Weiterhin konnten in der vorliegenden Studie zwei Bindungsstellen für das wichtigste Transportsubstrat, ATP, ermittelt werden. Eine dieser Bindungsstellen umfasst die N-terminale α-Helix, den Linker und die nahegelegenen β-Stränge. Die Lokalisierung der ATP-Bindestellen deutet kontrollierten Metabolitenfluss und durch den Liganden induzierte Stabilisierung des offenen Zustands der VDAC1-Pore an. Zum Abschluss zeigt diese Studie, dass Ca2+ mit zwei unterschiedlichen N- und C-terminalen Bereichen des β-Fasses interagiert. Diese Regionen überlappen mit Oligomerisierungsstellen und dynamischen Regionen von VDAC1 und weisen daher auf eine Verbindung zwischen Ca2+-Wechselwirkung, Kanalschließung und Oligomerisierung hin.The voltage-dependent anion channel
(VDAC), the most abundant protein in the outer mitochondrial
membrane, acts as a gatekeeper for the entry and exit of
mitochondrial metabolites and is involved in mitochondrial
apoptosis. Recent determination of its high-resolution structure by
three independent groups revealed a 19-stranded β-barrel with a
differently arranged N-terminal α-helix inside the pore. In this
thesis, the NMR resonance assignment of isoform one of human VDAC
(VDAC1) in solution is increased. In addition, this study provides
evidence for a kinked α-helical structure of the N-terminus, which
is compatible with the crystal structure of VDAC1, although other
structures cannot be excluded. Furthermore, this study reveals
functional dynamics of VDAC1 by a combination of solution NMR
spectroscopy, Gaussian network model (GNM) analysis, and molecular
dynamics (MD) simulations. Low signal intensities indicate
conformational exchange in the second part of the N-terminal
α-helix and the linker connecting the α-helix to the first
β-strand. In addition, mutation of arginine 15 in the second
α-helical part affects the stability of the α-helix and the overall
barrel in a complex manner. Micro- to millisecond dynamics are
significantly increased in the N-terminal α-helix, the linker, and
the N-terminal six β-strands of VDAC1 in micellar solution. In
addition, hydrogen bonds are instable in the N-terminal three
β-strands. In agreement, the N-terminal β-strands exhibit increased
B-factors in the crystal structure of VDAC1 and intrinsic
instability predicted by the GNM analysis. Mutation or chemical
modification of the membrane facing glutamic acid 73 (E73) strongly
reduces the micro- to millisecond dynamics in solution. MD
simulations reveal that a charge on E73 accounts for the elevation
of N-terminal protein dynamics as well as a thinning of the nearby
membrane. Since E73 is necessary for hexokinase-I-induced VDAC1
channel closure and inhibition of apoptosis, these results imply
that micro- to millisecond dynamics in the N-terminal part of the
β-barrel are essential for VDAC1 interaction and gating. Moreover,
the data suggest that dynamics in the α-helix are connected with
these processes. Furthermore, this study reveals two binding sites
for the pore's most important transport substrate ATP. The location
of the ATP binding sites, one of them comprising the N-terminal
α-helix, the linker, and nearby β-strands, suggests controlled
metabolite flux and ligand-induced stabilization of the open state
of the VDAC1 pore. Finally, Ca2+ is found to interact with two
distinct N-terminal and C-terminal regions in the β-barrel. These
regions overlap with VDAC1 oligomerization sites and dynamic
regions, suggesting a connection between Ca2+ interaction, gating,
and oligomerization.2013-02-2
Voltage Dependence of Conformational Dynamics and Subconducting States of VDAC-1
No full textAbstractThe voltage-dependent anion channel 1 (VDAC-1) is an important protein of the outer mitochondrial membrane that transports energy metabolites and is involved in apoptosis. The available structures of VDAC proteins show a wide β-stranded barrel pore, with its N-terminal α-helix (N-α) bound to its interior. Electrophysiology experiments revealed that voltage, its polarity, and membrane composition modulate VDAC currents. Experiments with VDAC-1 mutants identified amino acids that regulate the gating process. However, the mechanisms for how these factors regulate VDAC-1, and which changes they trigger in the channel, are still unknown. In this study, molecular dynamics simulations and single-channel experiments of VDAC-1 show agreement for the current-voltage relationships of an “open” channel and they also show several subconducting transient states that are more cation selective in the simulations. We observed voltage-dependent asymmetric distortions of the VDAC-1 barrel and the displacement of particular charged amino acids. We constructed conformational models of the protein voltage response and the pore changes that consistently explain the protein conformations observed at opposite voltage polarities, either in phosphatidylethanolamine or phosphatidylcholine membranes. The submicrosecond VDAC-1 voltage response shows intrinsic structural changes that explain the role of key gating amino acids and support some of the current gating hypotheses. These voltage-dependent protein changes include asymmetric barrel distortion, its interaction with the membrane, and significant displacement of N-α amino acids
Large-scale comparison of protein essential dynamics from molecular dynamics simulations and coarse-grained normal mode analyses
No full textSynthesis and Characterization of A Stable Non-cyclic Bis(amino)arsenium Cation
No full textThe reaction of Li[Mes*NH] (1, Mes * = 2,4,6-tri-tert-butylphenyl) with aminoarsane Mes*N(H)AsCl2 (2, Mes* = 2,4,6-t-Bu3C6H2) at –80 °C resulted in the formation of bisamino(chloro)arsane (Mes*NH)2AsCl (3Cl) by elimination of LiCl. 3Cl reacted with the Lewis acids such as AlCl3, GaCl3 and Ag[X] (X = AsF6–, OTf–, BF4–; OTf = trifluoromethanesulfonate = OSO2CF3–) upon chloride ion abstraction to give salts bearing the cation [(Mes*NH)2As]+ (3[X]; X = AsF6–, OTf–, BF4–, ECl4; E = Al, Ga). 3+ represents the first NH-functionalized acyclic bis(amino)arsenium cation. The formation of the salts bearing 3+ could also be observed in the reaction of cyclo-1,3-diarsa-2,4-diazane [ClAs(μ-NMes*)]2 (4) with Lewis acids (AlCl3, GaCl3) in the presence of proton sources in solution. All presented salts 3[X] were stable at room temperature and fully characterized.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author
