Outer Gate Research Articles

A Modular Heat-Sensing Domain Activates K2P2.1 through a Heat-Insensitive Gate

Ion channels often have modular structure with dedicated gating and sensory domains. Ligand binding to a localized site in the sensory domain usually affects channel function through allosteric effects on the gate. It is not clear whether this paradigm is applicable to ion channels regulated by temperature. Indeed, temperature may have an effect on the whole channel structure and thus, bypass the need for a dedicated heat-sensing domain. Heat leads to activation of the ‘leak’ two-pore (K2P) potassium channel K2P2.1 (KCNK2/TREK-1) through opening of a selectivity filter-based outer gate in the extracellular pore domain. We sought to investigate whether the pore plays a role in sensing heat or whether it only functions as a gate that responds to the commands from the temperature-sensing elements located elsewhere in the channel. We found that the incorporation of mutations designed to prevent allosteric communication between the intracellular C-terminal domain and the pore domain of K2P2.1 resulted in channels that remain functional but fail to respond to temperature. These results indicate that the gating mechanism of the pore lacks intrinsic temperature sensitivity and that the heat-sensing elements of K2P2.1 are confined within its C-terminus. Our data provide experimental support for the general notion of the existence of modular temperature sensing domains and highlight functional distinction between gating and sensory elements in heat-sensitive ion channels.

Insight into the Alternating Access Mechanism of the Sodium Symporter Mhp1 using Path Sampling Simulations

Sodium coupled co-transporters of the LeuT superfamily use an alternating access mechanism to move small molecules across the cell membrane. A key step in the transport cycle involves the transporter undergoing a conformational transition from a state poised to bind extracellular substrates to one that is competent to deliver substrate to the cytoplasm. To investigate this step in semi-atomistic detail, we construct a coarse-grained model of the benzylhydantoin transporter Mhp1 that incorporates experimental structures of the outward- and inward-open states. We obtain a rigorous sampling of the unbiased outward- to inward-facing transition path ensemble (TPE) using the weighted ensemble path-sampling method. Analysis of the TPE reveals two distinct sets of pathways connecting the inward- and outward-facing conformations. The first is consistent with a strict alternating access mechanism, while in the other, decoupling of the inner and outer gates transiently forms a putative permeation pathway through the transporter. Our simulations additionally show that transitions between the outward- and inward-open states result from rigid body motions of the hash motif relative to the substrate bundle, supporting the rocking bundle hypothesis. This methodology provides the groundwork for more chemically detailed investigations of the transport mechanism of superfamily members using fully atomistic simulations.

Enhancement of Closed-State Inactivation by Neutralization of S4 Arginines in Domain IV of a Sodium Channel

In PNAS, Capes et al. (2012) showed that the voltage sensor of domain IV in the skeletal muscle voltage-dependent sodium channel is energetically coupled to the channel gate located at the outer side of the pore. For this, the authors elicited gating pore currents, also known as omega currents, through each of the channel's four voltage sensors, one at a time. They inventively showed that sodium channel blockers TTX and μ-CTX, which bind to the outer pore in the selectivity filter region of the channel, affect the gating pore currents through domain IV, suggesting a coupling between conformational transitions in the selectivity filter and the movement of S4 in the voltage sensor of domain IV. The authors did not refer to two important aspects, which are tightly related to their experimental approach. The first is that neutralization of arginines on S4 (by R to Q substitutions) used to elicit gating pore currents through each of the voltage sensor mutants, may abolish the voltage sensitivity of the voltage sensor (Bao et al., 1999), and position it in the activated or partial activated state (Gagnon and Bezanilla, 2009). The second aspect relates to the predominant role that domain IV voltage sensor plays in coupling activation with fast inactivation in sodium channels. The voltage sensor of domain IV is specialized in energizing a cytoplasmic inactivation element (IFM), connecting the third and fourth homologous domains, into the inner mouth of the pore via a mechanism known as “ball and chain” fast inactivation. Further, it was shown that a neutralizing mutation at the outermost S4 arginine in domain IV, which is associated with the paramyotonia congenital disease, greatly affects the coupling between activation and inactivation in the rat and human skeletal muscle sodium channels (Chahine et al., 1994). It is therefore likely that activation of the voltage sensor of domain IV by the neutralizing mutations shifts the channel into a closed-state inactivation (Armstrong, 2006), which is associated with gating transitions that favor TTX binding to the outer pore (Patton and Goldin, 1991). If this is indeed the case, it points out to coupling between closed-state gating transitions at the outer pore gates with virtually all of the four voltage sensors, and not only with that of domain IV. Accelerated transitions in the selectivity filter at the outer gate due to occlusion of the cytoplasmic side of the pore by N-type inactivation particles is well established in homotetrameric K+ channels, though the parallel process in eukaryotic sodium channels is still under debate. The technical limitation arises by channel entrance into a closed-state gating transitions due to neutralization of S4 arginines should be addressed to refine the experimental approach used by Capes et al. in order to make it more reliable. For example, it is possible to bring each of the mutant channels into a uniform gating state using a conditioning protocol that imposes channel entrance into a closed-state inactivation prior to recording of the gating pore currents, or use a mutant channel in which its fast inactivation is removed by mutating the IFM motif between domains III to IV.

Simulations of the Alternating Access Mechanism of the Sodium Symporter Mhp1

Sodium coupled cotransporters of the five-helix inverted repeat (5HIR) superfamily use an alternating access mechanism to transport a myriad of small molecules across the cell membrane. One of the primary steps in this mechanism is the conformational transition from a state poised to bind extracellular substrates to a state that is competent to deliver substrate to the cytoplasm. Here, we construct a coarse-grained model of the 5HIR benzylhydantoin transporter Mhp1 that incorporates experimental structures of the outward- and inward-open states to investigate the mechanism of this conformational change. Using the weighted ensemble path-sampling method, we rigorously sample the outward- to inward-facing transition path ensemble. The transition path ensemble reveals a heterogeneous set of pathways connecting the two states and identifies two modes of transport: one consistent with a strict alternating access mechanism and another where decoupling of the inner and outer gates causes the transient formation of a continuous permeation pathway through the transporter. We also show that the conformational switch between the outward- and inward-open states results from rigid body motions of the hash motif relative to the substrate bundle, supporting the rocking bundle hypothesis. Finally, our methodology provides the groundwork for more chemically detailed investigations of the alternating mechanism.

Palytoxin action on the Na +,K +-ATPase and the disruption of ion equilibria in biological systems

Palytoxin-group toxins (PlTX) exert their potent biological activity by altering mechanisms of ion homeostasis in excitable and non-excitable tissues. This review will describe major aspects that led to the relatively early identification of the Na +,K +-ATPase as the molecular target and receptor of the toxin in sensitive systems. The importance of this pump in the normal functioning of animal cells has driven extensive investigative efforts. The recognized molecular mechanism of action of PlTX involves its binding to the extracellular portion of alpha subunit of this plasma membrane protein, which converts an enzyme carrying ions against their concentration gradients at the expense of chemical energy (ATP) into a non-selective cation channel, allowing passive flow of ions following their concentration gradients. More recent findings have indicated that PlTX would interfere with the normal strict coupling between inner and outer gates of the pump controlling the ion access to the Na +,K +-ATPase, allowing the gates to be simultaneously open. The ability of PlTX to make internal portions of the Na +,K +-ATPase accessible to relatively large molecules has been exploited to characterize the structure–function relationship of the pump, leading to a better understanding of its ion translocation pathway. Thus, forty years from the isolation of this potent marine biotoxin, a considerable understanding of its mode of action and of its potential as a research tool have been achieved and are the basis for promising future advancement in the characterization of biological systems and their alteration by PlTX.

Three‐dimensional flow simulation of resin transfer molding utilizing multilayered fiber preform

AbstractImpregnation of the multi‐layered fiber preform with injected resin was analyzed by using the control volume finite element method for three‐dimensional mold filling simulation of the resin transfer molding (RTM). Numerically predicted flow fronts were compared with experimental data to validate accuracy of the numerical results and applicability of the numerical code developed in this study. Since isothermal Newtonian flow was assumed, Darcy's law and continuity equation were used as governing equations and permeability tensors employed for each layer. Flow simulation was conducted for the two types of mold geometries, i.e., rectangular plate and hollow cylinder. It was proven by comparing numerical results with experimental data that the simulation code is accurate enough to predict the flow patterns, especially when multi‐layered preforms were placed in the cylindrical mold. It was also found that the filling time was reduced by using outer gates compared with the case of inner gates. Three‐dimensional numerical simulation provided useful information for mold filling and can be used to design an optimum mold and make the RTM process efficient. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009

Gating the pore of potassium leak channels

A key feature of potassium channel function is the ability to switch between conducting and non-conducting states by undergoing conformational changes in response to cellular or extracellular signals. Such switching is facilitated by the mechanical coupling of gating domain movements to pore opening and closing. Two-pore domain potassium channels (K(2P)) conduct leak or background potassium-selective currents that are mostly time- and voltage-independent. These channels play a significant role in setting the cell resting membrane potential and, therefore modulate cell responsiveness and excitability. Thus, K(2P) channels are key players in numerous physiological processes and were recently shown to also be involved in human pathologies. It is well established that K(2P) channel conductance, open probability and cell surface expression are significantly modulated by various physical and chemical stimuli. However, in understanding how such signals are translated into conformational changes that open or close the channels gate, there remain more open questions than answers. A growing line of evidence suggests that the outer pore area assumes a critical role in gating K(2P) channels, in a manner reminiscent of C-type inactivation of voltage-gated potassium channels. In some K(2P) channels, this gating mechanism is facilitated in response to external pH levels. Recently, it was suggested that K(2P) channels also possess a lower activation gate that is positively coupled to the outer pore gate. The purpose of this review is to present an up-to-date summary of research describing the conformational changes and gating events that take place at the K(2P) channel ion-conducting pathway during the channel regulation.

Two Distinct Molecular Mechanisms Underlie pH Sensitivity of the Human Potassium Leak Channel K2P2.1

The mammalian K2P2.1 potassium channel (TREK-1, KCNK2) is highly expressed in excitable tissues, where it plays a key role in the cellular mechanisms of neuroprotection, anaesthesia, pain perception and depression. Here, we report that external acidification, within the physiological range, strongly inhibits the human K2P2.1 channel in two distinct time scales. We have identified two histidine residues (i.e., H87 and H141), located in the first external loop of the channel, which govern the fast response of the channel to external pH (in the time scale of seconds). We demonstrate that these residues are within physical proximity to glutamate 84, homologous to Shaker E418, KcsA E51 and KCNK0 E28 residues, all previously reported to stabilize the outer pore gate in the open conformation by forming hydrogen bonds with pore-adjacent residues. We thus propose a novel mechanism for pH sensing in which protonation of H141 and H87 generates a local positive charge that serves to draw E84 away from its natural interactions, facilitating the collapse of the selectivity filter region, a mechanism which resembles C-type gating of voltage dependent potassium channels. In accordance with this proposed mechanism, the proton-mediated effect was inhibited by external potassium ions, modified the channel's ion selectivity and was enhanced by a mutation, S164Y, known to accelerate C-type gating. In addition, we show that the slow regulatory effect (in the time scale of minutes) is mediated by proton-sensitive G-protein coupled receptors (GPCRs), which activated phospholipase C via the Gq pathway. We demonstrate that three residues within the C-terminal of K2P2.1 mediate the channel's response to the observed GPCRs activation by acidic pH.Taken together, our results highlight the physiological importance of human K2P2.1 channels as sensors of extracellular pH in the central nervous system.

Distinct gate conformations of the ABC transporter BtuCD revealed by electron spin resonance spectroscopy and chemical cross-linking

BtuCD is a type II ABC importer that catalyzes the translocation of vitamin B12 from the periplasm into the cytoplasm of Escherichia coli. Crystal structures of BtuCD and the related HiF (or Hi1470/71) protein from Haemophilus influenzae have revealed distinct conformations of the transmembrane domains that form inner and outer gates. We used electron spin resonance spectroscopy to study the reaction cycle of BtuCD after labeling the protein at residues located at these gates. The results suggest that BtuCD as a prototype type II ABC importer may have a mechanism that is distinct from that of ABC exporters such as Sav1866 or type I ABC importers such as those specific for molybdate (ModBC) or maltose (MalFGK). Structured summary MINT- 6803800: btuF (uniprotkb: P37028), btuC (uniprotkb: P06609) and btuD (uniprotkb: P06611) physically interact (MI: 0218) by molecular sieving (MI: 0071)

A Novel Mechanism for Human K2P2.1 Channel Gating: FACILITATION OF C-TYPE GATING BY PROTONATION OF EXTRACELLULAR HISTIDINE RESIDUES

The mammalian K2P2.1 potassium channel (TREK-1, KCNK2) is highly expressed in excitable tissues, where it plays a key role in the cellular mechanisms of neuroprotection, anesthesia, pain perception, and depression. Here, we report that external acidification, within the physiological range, strongly inhibits the human K2P2.1 channel by inducing "C-type" closure. We have identified two histidine residues (i.e. His-87 and His-141), located in the first external loop of the channel, that govern the response of the channel to external pH. We demonstrate that these residues are within physical proximity to glutamate 84, homologous to Shaker Glu-418, KcsA Glu-51, and KCNK0 Glu-28 residues, all previously argued to stabilize the outer pore gate in the open conformation by forming hydrogen bonds with pore-adjacent residues. We thus propose a novel mechanism for pH sensing in which protonation of His-141 and His-87 generates a local positive charge that serves to draw Glu-84 away from its natural interactions, facilitating the collapse of the selectivity filter region. In accordance with this proposed mechanism, low pH modified K2P2.1 selectivity toward potassium. Moreover, the proton-mediated effect was inhibited by external potassium ions and was enhanced by a mutation (S164Y) known to accelerate C-type gating. Furthermore, proton-induced current inhibition was more pronounced at negative potentials. Thus, voltage-dependent C-type gating acceleration by protons represents a novel mechanism for K2P2.1 outward rectification.

Jena Romanticism and Zen

Jena Romanticism and Zen Dennis McCort (bio) But what drew him with all its power was a tall, light blue flower that stood beside the stream and touched him with its broad, glittering petals. Surrounding it were countless flowers of all colors, and the rare fragrance filled the air. He saw nothing but the blue flower and regarded it for a long while with indescribable tenderness. Finally he made as if to approach it when it suddenly stirred and began to change. The petals became even more brilliant and pressed themselves against the growing stem. The flower inclined towards him, its petals forming a broad blue collar in which a tender face hovered. —Novalis, Heinrich von Ofterdingen1 All at once, his biographer relates, Bassui felt as though he had "lost his life root, like a barrel whose bottom had been smashed open." Sweat began to pour from every pore of his body, and when he left Koho's room he was in such a daze that he bumped his head several times along the walls trying to find the outer gate of the temple. Upon reaching his hut he wept for hours from his very depths. The tears overflowed, "pouring [End Page 98] down his face like rain." In the intense combustion of this overwhelming experience Bassui's previously held conceptions and beliefs, we are told, were utterly annihilated. —Kapleau, The Three Pillars of Zen Introduction Two scenarios above: the first presents the climax of a lucid dream in which Heinrich, the poet-hero of Novalis' signature novel of German romanticism, experiences an apotheosis of nature centering on the mystical Blue Flower, a mandala whose dynamic perfection of rounded form reveals to him the ecstatic unity of all being. In the second, the medieval Rinzai Zen monk Bassui undergoes the near-violent collapse of an ego-based personality-structure eroded by years of meditation and the consequent influx of an all-pervasive radiance. Though different in particulars (including the difference between fiction and biography), both texts convey the same apocalyptic, life-transforming event, known to Zen as satori and to German romanticism, variously, as die Erleuchtung, romantisieren, or das goldene Zeitalter. This is the summum bonum sought by both these spiritual movements, the goal of all spiritual practice, intellectual inquiry and emotional struggle, the pearl of great price worthy of any sacrifice. It brings with it an absolute freedom empowering one to see anything in one's field of consciousness exactly as it is in itself. It reveals nothing less than the noumenal Ding an sich of Kantian transcendentalism, or, as Zen puts it, things in their exquisite "suchness" (Skt., tathatã). Let us refer to this event in the ensuing discussion as "enlightenment," the term generally used by Westerners to signify the supreme goal of Hindu and Buddhist spiritual practice. It is an apt term inasmuch as many, if not most, such experiences are characterized by those who have them as suffused with a profound sense of light or radiance, albeit a radiance in no way oppressive to one's physical vision. Philip Kapleau, an American Zen master, equates enlightenment with "Self-realization," probably both in Jung's sense of individuation (the maturation and harmonization of psychic forces) and in that of the early Chinese master Hui-neng who characterized the event as "seeing into one's own True Nature." Kapleau goes on to describe enlightenment as "opening the Mind's eye, awakening to one's True-nature and hence of [sic] the nature of all existence" (377). [End Page 99] This much description should be sufficient to convey a sense of enlightenment as it is understood by its devotees, that is, as the ultimate emancipation of human consciousness from the prison of separateness and the pressure of any and all constraint, internal or external. This makes both Jena romanticism and Zen an "argument" in favor of the possibility of "pure" or "direct" experience, of consciousness unfettered by the barest taint of cultural conditioning. Both Novalis and Hui-neng insist that there is such a thing as transcendence and that it is "there for the taking," given the requisite surrender of narrow self-interest. The above snippet of Bassui...

Between Human Rights and the Rule of Law: Indefinite Detention and the Derogation Model of Constitutionalism

The attacks on America on September 11, 2001 heralded an era of what has been described as defensive democracy.' Governments across the democratic world have acted to concentrate power into their own hands whilst shutting their outer gates. But although strength is undoubtedly the first necessity of any constitution, the line between strength and unbridled power is dangerously fine. Moreover, the first casualties of defensive measures are inevitably the civil liberties and human rights that individuals normally enjoy. It is therefore imperative that in order to maintain the very democratic systems that defensive postures are intended to protect, state constitutions operate to defend the threatened bridgeheads of legality and human rights. This issue is raised in no more stark form than in relation to the UK's postSeptember 11 derogation from the European Convention on Human Rights and the resistance to derogation (as well as to continued derogation) that has been provided by the non-governmental institutions of the UK's constitutional regime. The extent of the judicial resistance to derogation has now been authoritatively established by the House of Lords in Av. Secretary of Statefor the Home Department.2 This paper examines the decision against the background of three differing constitutional models relating to the nature of derogation. Each model represents a very different approach to the defence of the twin bridgeheads of legality and human rights. These models offer a perspective from which to evaluate the House of Lords' decision and, to some degree, we find each reflected in the decision. But before turning to these models and to the critical analysis of the decision, it is useful to begin by setting out the ruling and its immediate context.

Fractionation of Alfalfa into Leaves and Stems using a Three Pass Rotary Drum Dryer

Fractionation/separation of chopped alfalfa into leaf and stem components was investigated using a small industrial three-pass rotary drum dryer. Both drying and fractionation of chopped alfalfa were successfully achieved in a single operational process. Generally, fractionation is carried out only as a separate process and, therefore, more costly overall. The success of fractionation was determined by the total amount of the desired component (leaf or stem) recovered, relative to the amount entering the process (i.e. separation efficiency) and by the purity (leaf or stem proportion) of the component in a product stream collected at the cyclone or drum exit. A series of experiments under different operating conditions were performed to establish the optimum process conditions leading to maximum separation efficiency and leaf purity. The operating variables used during experimentation were the outer drum end gate opening area, material feed rate, gas flow rate and gas temperatures at the drum inlet. The experimental results indicated that leaf purity decreased and separation efficiency increased with a decrease in dryer drum end gate opening area. A decrease in separation efficiency was observed with an increase in material feed rate. On the other hand, leaf purity increased and separation efficiency decreased with a decrease in gas flow rate. The decrease in separation efficiency was most significant at low gas flow rates. Separation efficiency decreased with a decrease in gas inlet temperature. Stem moisture content was observed to be consistently higher than the leaves. Lowering stem moisture levels to an acceptable level of 8% moisture content results in over drying of the leaves, leading to a loss in product quality. It was determined that the relative difference in moisture levels of stems and leaves at the drum and cyclone exit, respectively, can be averted by re-circulating in the order of 20% of the wet material (stems) from the drum exit back through the dryer inlet.

日比谷公園開園時における二・三の施設の石材加工・利用技術

This study examined into the technology of working and utilization of stone for some facilities, (1) entrance gateposts, (2) roadway curbstones and (3) pond bank stones in the beginning Hibiya Park (1903, Tokyo) by field survey and literature research. (1) The gateposts of Hibiya Park's 6 entrances were similar to gateposts of western-style building's exterior, and those were made by Japanese stonemasonry. The stones of gateposts were reused andesite or granite stones of the walls with outer castle gates of Edo castle. (2) The Hibiya park's western-style roadway set granite curbstones and concrete gutter. And the pedestrian way side in the park was set cobblestone curbstones and gutter. The cobblestones were local gardening material in Edo. (3) The bank of Shinji-ike-pond (Shinji means Chinese character 'Kokoro') : Japanese-style pond laid cobblestones randomly like the ponds of gardens in Edo. The bank of Kumogata-ike-pond (Kumogata means 'French curve') : western-style pond laid single sized cobblestones for artificial shape. Then, technology of working and utilization of stone for some facilities in the beginning Hibiya park were based on western style architecture and civil engineering, and exploited Japanese technique - skills and materials of Edo.

Potassium-Dependent Slow Inactivation of Kir1.1 (ROMK) Channels

The ROMK (Kir1.1) family of epithelial K channels can be inactivated by a combination of low internal pH and low external K, such that alkalization does not reopen the channels unless external K is elevated. Previous work suggested that this inactivation results from an allosteric interaction between an inner pH gate and an outer K sensor, and could be described by a simple three-state kinetic model. In the present study, we report that a sustained depolarization slowly inactivated (half-time = 10–15 min) ROMK channels that had been engineered for increased affinity to internal polyamines. Furthermore, this inactivation occurred at external [K] ≤1 mM in ROMK mutants whose inner pH gate was constitutively open (ROMK2-K61M mutation). Both pH and voltage inactivation depended on external K in a manner reminiscent of C-type inactivation, but having a much slower time course. Replacement of ROMK extracellular loop residues by Kir2.1 homologous residues attenuated or abolished this inactivation. These results are consistent with the hypothesis that there are (at least) two separate closure processes in these channels: an inner pH-regulated gate, and an outer (inactivation) gate, where the latter is modulated by both voltage and external [K].

Ion occlusion/deocclusion partial reactions in individual palytoxin-modified Na/K pumps.

In P-type ion-motive ATPases, transported ions approach their binding sites from one membrane surface, become buried deep within "occluded" conformations in which the sites are inaccessible from either membrane side, and are then deoccluded and released to the opposite membrane surface. This describes an alternating-gate transport mechanism, in which the pump acts like an ion channel with two gates that open and close alternately. The occluded states ensure that one gate closes before the other can open, thus preventing the large electrodiffusive ion fluxes that would otherwise quickly undo the pump's electrochemical work. High-resolution crystal structures of two conformations of the SERCA (sarcoplasmic and endoplasmic reticulum Ca(2+)) P-type ATPase, together with mutagenesis results and analyses of structural models based on homology, have begun to provide a picture of the ion coordination sites in related P-type ATPases, including the Na/K pump. However, in no P-type ATPase are the structures and mechanisms of the gates known. The marine toxin, palytoxin (PTX), is known to bind to the Na/K pump and elicit a nonselective cation leak pathway, possibly by disrupting the strict coupling between the pump's inner and outer gates, allowing them to both be open. We recently found that ion flow through PTX-modified Na/K pump-channels appears to be modulated by two gates that can be regulated by the pump's physiological ligands in a manner suggesting that gating reflects underlying ion occlusion/deocclusion partial reactions. We review that work here and provide evidence that the pore of the PTX-induced pump-channel has a diameter > 6 A.

Early Fluorescence Signals Detect Transitions at Mammalian Serotonin Transporters

The mammalian serotonin transporters rSERT or hSERT were expressed in oocytes and labeled with sulforhodamine-MTS. The endogenous Cys-109 residue contributes most of the signal, and the labeled transporter shows normal function. The SERT fluorescence decreases in the presence of 5-HT and also depends on the inorganic substrates of SERT. The fluorescence also increases with membrane depolarization. During voltage-jump experiments, fluorescence relaxations show little inactivation or history dependence. The fluorescence signal has a voltage dependence similar to that of the prepriming step of the previously described voltage-dependent transient current. However, the fluorescence relaxations are the fastest voltage-dependent events yet studied at SERT; their time constants of ∼8–30ms are severalfold faster than the prepriming or inactivation phases of the transient currents. These fluorescence signals are interpreted within the framework of the gate-lumen-gate model. The signals may monitor initial events at the outer gate.

KCNKØ: Opening and Closing the 2-P-Domain Potassium Leak Channel Entails “C-Type” Gating of the Outer Pore

Essential to nerve and muscle function, little is known about how potassium leak channels operate. KCNKØ opens and closes in a kinase-dependent fashion. Here, the transition is shown to correspond to changes in the outer aspect of the ion conduction pore. Voltage-gated potassium (VGK) channels open and close via an internal gate; however, they also have an outer pore gate that produces “C-type” inactivation. While KCNKØ does not inactivate, KCNKØ and VGK channels respond in like manner to outer pore blockers, potassium, mutations, and chemical modifiers. Structural relatedness is confirmed: VGK residues that come close during C-type gating predict KCNKØ sites that crosslink (after mutation to cysteine) to yield channels controlled by reduction and oxidization. We conclude that similar outer pore gates mediate KCNKØ opening and closing and VGK channel C-type inactivation despite their divergent structures and physiological roles.

Self-similar conductance fluctuations in a Sinai billiard with a mixed chaotic phase space

We have investigated ballistic transport through a sub-micron-sized Sinai billiard fabricated using a high-mobility two-dimensional electron gas. The geometry of the billiard is defined by a circular surface gate surrounded by an outer square gate with two quantum point contacts. When both gates are biased negatively, the Sinai billiard is formed and collisions with the confining potential walls generate chaotic classical motion. Recent experiments have revealed novel ‘self-similar’ fluctuation patterns in the conductance of the Sinai billiard, measured as a function of magnetic field perpendicular to the plane of the electron gas. We have undertaken quantum mechanical calculations of the device conductance using a detailed model for the potential landscape under the surface gates. We compare these results with predictions for a hard-walled billiard and show that the surface gate geometry can have a large effect on the observed transport properties.

Essential Role of Cooperative Swing of Loops in the Transport Function of Mitochondrial ADP/ATP Carrier.

The mitochondrial ADP/ATP carrier, which mediates transport of ADP and ATP via the inner membrane, functions as a dimer, each subunit being located in the same orientation. The carrier has three large loops (M1, M2, M3) facing the matrix side. M1 loop is suggested to function as an outer gate, M2 as a substrate binding site and M3 as an auxiliary gate by interacting with other loops. The transport function of the carrier is characterized largely by the cooperative swing of these paired loops, in which location of loops changes repeatedly between outside and inside of the membrane.