Voltage-dependent Gating Research Articles

Electromechanical coupling mechanism for activation and inactivation of an HCN channel

Pacemaker hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channels exhibit a reversed voltage-dependent gating, activating by membrane hyperpolarization instead of depolarization. Sea urchin HCN (spHCN) channels also undergo inactivation with hyperpolarization which occurs only in the absence of cyclic nucleotide. Here we applied transition metal ion FRET, patch-clamp fluorometry and Rosetta modeling to measure differences in the structural rearrangements between activation and inactivation of spHCN channels. We found that removing cAMP produced a largely rigid-body rotation of the C-linker relative to the transmembrane domain, bringing the A’ helix of the C-linker in close proximity to the voltage-sensing S4 helix. In addition, rotation of the C-linker was elicited by hyperpolarization in the absence but not the presence of cAMP. These results suggest that — in contrast to electromechanical coupling for channel activation — the A’ helix serves to couple the S4-helix movement for channel inactivation, which is likely a conserved mechanism for CNBD-family channels.

Connexin 46 and connexin 50 gap junction channel properties are shaped by structural and dynamic features of their N-terminal domains.

Gap junctions formed by different connexins are expressed throughout the body and harbour unique channel properties that have not been fully defined mechanistically. Recent structural studies by cryo-electron microscopy have produced high-resolution models of the related but functionally distinct lens connexins (Cx50 and Cx46) captured in a stable open state, opening the door for structure-function comparison. Here, we conducted comparative molecular dynamics simulation and electrophysiology studies to dissect the isoform-specific differences in Cx46 and Cx50 intercellular channel function. We show that key determinants Cx46 and Cx50 gap junction channel open stability and unitary conductance are shaped by structural and dynamic features of their N-terminal domains, in particular the residue at the 9th position and differences in hydrophobic anchoring sites. The results of this study establish the open state Cx46/50 structural models as archetypes for structure-function studies targeted at elucidating the mechanism of gap junction channels and the molecular basis of disease-causing variants. Connexins form intercellular communication channels, known as gap junctions (GJs), that facilitate diverse physiological roles, from long-range electrical and chemical coupling to coordinating development and nutrient exchange. GJs formed by different connexin isoforms harbour unique channel properties that have not been fully defined mechanistically. Recent structural studies on Cx46 and Cx50 defined a novel and stable open state and implicated the amino-terminal (NT) domain as a major contributor for isoform-specific functional differences between these closely related lens connexins. To better understand these differences, we constructed models corresponding to wildtype Cx50 and Cx46 GJs, NT domain swapped chimeras, and point variants at the 9th residue for comparative molecular dynamics (MD) simulation and electrophysiology studies. All constructs formed functional GJ channels, except the chimeric Cx46-50NT variant, which correlated with an introduced steric clash and increased dynamical behaviour (instability) of the NT domain observed by MD simulation. Single channel conductance correlated well with free-energy landscapes predicted by MD, but resulted in a surprisingly greater degree of effect. Additionally, we observed significant effects on transjunctional voltage-dependent gating (Vj gating) and/or open state dwell times induced by the designed NT domain variants. Together, these studies indicate intra- and inter-subunit interactions involving both hydrophobic and charged residues within the NT domains of Cx46 and Cx50 play important roles in defining GJ open state stability and single channel conductance, and establish the open state Cx46/50 structural models as archetypes for structure-function studies targeted at elucidating GJ channel mechanisms and the molecular basis of cataract-linked connexin variants.

Dynamic Response of Arterial Cerebral Ca V 1.2 Channels to Intravascular Pressure

Intravascular pressure elicits arterial constriction by elevating intracellular [Ca2+] in vascular smooth muscle cells (SMCs). Past studies have argued that this rise is singularly linked to depolarization and the voltage-dependent gating of L-type (CaV1.2) Ca2+ channels. While important, pressure might also directly modulate CaV1.2 activity by promoting functional coupling among subunits, a response that enhances Ca2+ sparklets generation. In this regard, the study goal was to define the mechanosensitivity of CaV1.2 channels using a range of electrophysiological approaches. Beginning with whole-cell patch clamp electrophysiology, we show that SMCs stretching by a hypoosmotic challenge (from 300 to 200 mOsm) leads to a marked rise in total Ca2+ current (around 40%) without changing voltage-dependent activation/inactivation properties. This mechanosensitive response reflects augmented functional coupling, as revealed through recordings of single channel activity. Ongoing work has further revealed that the rise in functional coupling was intimately linked to caveolin-1, caveolae forming membrane protein tied to the cytoskeleton dynamics and signal transduction. This protein-protein relationship was also observed at the immunohistochemical level, with the proximity ligation assay showing that CaV1.2 colocalizes with caveolin-1 when SMCs are stretched by the hypoosmotic challenge. In closing, the current work reveals that CaV1.2 channels are a dynamic target of intravascular pressure and likely integral to a robust myogenic response. Continuing work will define the composition of the pressure-sensitive signaling complex and how it is impacted by disease.

High-Performance CVD Bilayer MoS2 Radio Frequency Transistors and Gigahertz Mixers for Flexible Nanoelectronics.

Two-dimensional (2D) MoS2 have attracted tremendous attention due to their potential applications in future flexible high-frequency electronics. Bilayer MoS2 exhibits the advantages of carrier mobility when compared with monolayer mobility, thus making the former more suitable for use in future flexible high-frequency electronics. However, there are fewer systematical studies of chemical vapor deposition (CVD) bilayer MoS2 radiofrequency (RF) transistors on flexible polyimide substrates. In this work, CVD bilayer MoS2 RF transistors on flexible substrates with different gate lengths and gigahertz flexible frequency mixers were constructed and systematically studied. The extrinsic cutoff frequency (fT) and maximum oscillation frequency (fmax) increased with reducing gate lengths. From transistors with a gate length of 0.3 μm, we demonstrated an extrinsic fT of 4 GHz and fmax of 10 GHz. Furthermore, statistical analysis of 14 flexible MoS2 RF transistors is presented in this work. The study of a flexible mixer demonstrates the dependence of conversion gain versus gate voltage, LO power and input signal frequency. These results present the potential of CVD bilayer MoS2 for future flexible high-frequency electronics.

Electron-electron interactions and weak antilocalization in few-layer ZrTe5 devices

Much effort has been devoted to the electronic properties of relatively thick ZrTe5 crystals, focusing on their three-dimensional topological effects. Thin ZrTe5 crystals, on the other hand, were much less explored experimentally. Here we present detailed magnetotransport studies of few-layer ZrTe5 devices, in which electron-electron interactions and weak anti-localization are observed. The coexistence of the two effects manifests themselves in corroborating evidence presented in the temperature and magnetic field dependence of the resistance. Notably, the temperature-dependent phase coherence length extracted from weak anti-localization agrees with strong electron-electron scattering in the sample. Meanwhile, universal conductance fluctuations have temperature and gate voltage dependence that is similar to that of the phase coherence length. Lastly, all the transport properties in thin ZrTe5 crystals show strong two-dimensional characteristics. Our results provide new insight into the highly intricate properties of topological material ZrTe5.

Interfacial symmetry breaking induced spin-orbit coupling in wurtzite GaN nanowires

The interfacial asymmetry-induced spin–orbit coupling (SOC) has a distinct effect on the spin relaxation and gate manipulation of spintronic devices. However, it is hard to separate the contributions from the interface and bulk by commonly used techniques, such as weak antilocalization and time-resolved Kerr rotation. Owing to the different distribution of the effective magnetic field in the momentum space, the SOC originated from the interfacial effect and the wurtzite structure inversion asymmetry (WSIA) could be distinguished by circular photo-galvanic effect (CPGE) measurements. In this work, the angle and external gate-voltage dependences of the interfacial SOC in GaN nanowires (NWs) were investigated using the CPGE. The interfacial SOC-induced CPGE shows a twofold degeneracy, while the removal of the sixfold degeneracy is attributed to the asymmetrical optical absorption. Furthermore, the interfacial SOC-induced CPGE shows a weak dependence on the external gate voltage, which is totally different from the WSIA-related SOC. This result is consistent with the clarification that the interfacial asymmetry originates from the Fermi level pinning at the GaN NW sidewall surfaces, thus clearly illustrating the special distribution and gate dependences of the interfacial SOC, which is critical for the design of spintronic devices based on GaN NWs.

Artificial Sub-60 Millivolts/Decade Switching in a Metal-Insulator-Metal-Insulator-Semiconductor Transistor without a Ferroelectric Component.

Negative capacitance field-effect transistors (NC-FETs) have attracted wide interest as promising candidates for steep-slope devices, and sub-60 millivolts/decade (mV/decade) switching has been demonstrated in NC-FETs with various device structures and material systems. However, the detailed mechanisms of the observed steep-slope switching in some of these experiments are under intense debate. Here we show that sub-60 mV/decade switching can be observed in a WS2 transistor with a metal-insulator-metal-insulator-semiconductor (MIMIS) structure without any ferroelectric component. This structure resembles an NC-FET with internal gate, except that the ferroelectric layer is replaced by a leaky dielectric layer. Through simulations of the charging dynamics during the device characterization using a resistor-capacitor network model, we show that the observed steep-slope switching in our "ferroelectric-free" transistors can be attributed to the internal gate voltage response to the chosen varying gate voltage scan rates. We further show that a constant gate voltage scan rate can also lead to transient sub-60 mV/decade switching in an MIMIS structure with voltage-dependent internal gate capacitance. Our results indicate that the observation of sub-60 mV/decade switching alone is not sufficient evidence for the successful demonstration of a true steep-slope switching device and that experimentalists need to critically assess their measurement setups to avoid measurement-related artifacts.

Spin-states in MoS2 thin-film transistors distinguished by operando electron spin resonance

Transition metal dichalcogenide MoS2 is a two-dimensional material, attracting much attention for next-generation applications thanks to rich functionalities stemming from its crystal structure. Many experimental and theoretical works have focused on the spin-orbit interaction which couples the valley and spin degrees of freedom so that the spin-states can be electrically controllable. However, the spin-states of charge carriers and atomic vacancies in devices have not been yet elucidated directly from a microscopic viewpoint. Here, we report the spin-states in thin-film transistors using operando electron spin resonance spectroscopy. We have observed clearly different electron spin resonance signals of the conduction electrons and atomic vacancies, and distinguished the corresponding spin-states from the signals and theoretical calculations, evaluating the gate-voltage dependence and the spin-susceptibility and g-factor temperature dependence. This analysis gives deep insight into the MoS2 magnetism and clearly indicates different spin-scattering mechanisms compared to graphene, which will be useful for improvements of the device characteristics and new applications.

Evidence of the Ferroelectric Polarization in Charge Transport through WTe2 Weyl Semimetal Surface

We investigate electron transport along the surface of WTe$_2$ three-dimensional single crystals, which are characterized by coexistence of Weyl semimetal conductivity and ferroelectricity at room temperature. We find that non-linear behavior of $dV/dI(I)$ WTe$_2$ differential resistance is accompanied by slow relaxation process, which appears as the $dV/dI(I)$ dependence on the sign of the current change. This observation is confirmed by direct investigation of time-dependent relaxation curves. While strongly non-linear differential resistance should be expected for the zero-gap WTe$_2$, the slow relaxation in transport is very unusual for well-conducting semimetals at room temperature. We establish, that non-monotonous dependence of the amplitude of the effect on driving current $\Delta dV/dI(I)$ well corresponds to the known Sawyer-Tower's ferroelectric hysteresis loop. This conclusion is also confirmed by gate voltage dependencies, so our results can be understood as a direct demonstration of WTe$_2$ ferroelectric polarization in charge transport experiment.

Structural mechanisms of gating and selectivity of human rod CNGA1 channel

Mammalian cyclic nucleotide-gated (CNG) channels play an essential role in the signal transduction of the visual and olfactory sensory systems. Here we reveal the structural mechanism of ligand gating in human rod CNGA1 channel by determining its cryo-EM structures in both the apo closed and cGMP-bound open states. Distinct from most other members of voltage-gated tetrameric cation channels, CNGA1 forms a central channel gate in the middle of the membrane, occluding the central cavity. Structural analyses of ion binding profiles in the selectivity filters of the wild-type channel and the E365Q filter mutant allow us to unambiguously define the two Ca2+ binding sites inside the selectivity filter, providing structural insights into Ca2+ blockage and permeation in CNG channels. The structure of the E365Q mutant also reveals two alternative side-chain conformations at Q365, providing a plausible explanation for the voltage-dependent gating of CNG channel acquired upon E365 mutation.

Epitaxial Al-InAs Heterostructures as Platform for Josephson Junction Field-Effect Transistor Logic Devices

We fabricate Josephson junction field-effect transistors (JJ-FETs) using InAs quantum well heterostructure as the channel, epitaxial Al as superconducting electrodes, and scaled-down Al2O3 gate dielectrics. A systematic investigation of the gate voltage ( ${V}_{{G}}$ ) dependence of the critical current, normal state conductance, and characteristic voltage of the JJ-FETs coupled with capacitance measurements reveals different ${V}_{{G}}$ regimes in the proximity effect characterization of JJ-FETs. Self-consistent Poisson–Schrodinger simulations allow us to associate these ${V}_{{G}}$ regimes with the carriers populating one or more subbands at different vertical locations in the heterostructure. We also discuss the importance of oxide/channel interface quality and its impact on the practical implementation of JJ-FET Boolean logic gates with signal restoration.

Gate voltage dependence of noise distribution in radio-frequency reflectometry in gallium arsenide quantum dots

We investigate gate voltage dependence of electrical readout noise in high-speed rf reflectometry using gallium arsenide quantum dots. The fast Fourier transform spectrum from the real time measurement reflects build-in device noise and circuit noise including the resonator and the amplifier. We separate their noise spectral components by model analysis. Detail of gate voltage dependence of the flicker noise is investigated and compared to the charge sensor sensitivity. We point out that the dominant component of the readout noise changes by the measurement integration time.

Magnetoelectric Response of Antiferromagnetic CrI3 Bilayers.

We predict that layer antiferromagnetic bilayers formed from van der Waals (vdW) materials with weak interlayer versus intralayer exchange coupling have strong magnetoelectric response that can be detected in dual-gated devices where internal displacement fields and carrier densities can be varied independently. We illustrate this strong temperature-dependent magnetoelectric response in bilayer CrI3 at charge neutrality by calculating the gate voltage-dependent total magnetization through Monte Carlo simulations and mean-field solutions of the anisotropic Heisenberg model informed from density functional theory and experimental data and present a simple model for electrical control of magnetism by electrostatic doping.

Approaching barrier-free contacts to monolayer MoS2 employing [Co/Pt] multilayer electrodes

The broken inversion symmetry and time-reversal symmetry along with the large spin–orbit interactions in monolayer MoS2 make it an ideal candidate for novel valleytronic applications. However, the realization of efficient spin-valley-controlled devices demands the integration of perpendicular magnetic anisotropy (PMA) electrodes with negligible Schottky barriers. Here, as the first demonstration, we fabricated a monolayer MoS2 field-effect transistor with PMA electrodes: Pt/[Co/Pt]3 and [Co/Pt]2. The I–V curves of PMA/MoS2 contacts show symmetric and linear behavior reflecting Ohmic nature. The flat-band Schottky barrier heights (SBHs) extracted using the temperature and gate voltage dependence of the I–V curves were found to be 10.2 and 9.6 meV. The observed SBHs are record low values reported thus far for any metal/monolayer MoS2 contact. High-quality PMA electrodes with almost zero SBH play a paramount role in the future development of novel spintronic/valleytronic devices; hence, our results can open a new route toward the realization of novel technological devices employing two-dimensional materials.

Inhibitory Mechanism of the Isoflavone Derivative Genistein in the Human CaV3.3 Channel.

Regulation of cellular excitability and oscillatory behavior of resting membrane potential in nerve cells are largely mediated by the low-voltage activated T-type calcium channels. This calcium channel family is constituted by three isoforms, namely, CaV3.1, CaV3.2, and CaV3.3, that are largely distributed in the nervous system and other parts of the body. Dysfunction of T-type calcium channels is associated with a wide range of pathophysiologies including epilepsy, neuropathic pain, cardiac problems, and major depressive disorders. Due to their pharmacological relevance, finding molecular agents able to modulate the channel's function may provide therapeutic means to ameliorate their related disorders. Here we used electrophysiological experiments to show that genistein, a canonical tyrosine kinase inhibitor, reduces the activity of the human CaV3.3 channel in a concentration-dependent manner. The inhibitory effect of genistein is independent of tyrosine kinase modulation and does not affect the voltage-dependent gating of the channel. Subsequently, we used computational methods to identify plausible molecular poses for the interaction of genistein and the CaV3.3 channel. Starting from different molecular poses, we carried out all-atom molecular dynamics (MD) simulations to identify the interacting determinants for the CaV3.3/genistein complex formation. Our extensive (microsecond-length) simulations suggest specific binding interactions that seem to stabilize the protein/inhibitor complex. Furthermore, our results from the unbiased MD simulations are in good agreement with the recently solved cryoelectron microscopy structure of the CaV3.1/Z944 complex in terms of both the location of the ligand binding site and the role of several equivalent amino acid residues. Proposed interacting complex loci were subsequently tested and corroborated by electrophysiological experiments using another naturally occurring isoflavone derivative, daidzein. Thus, by using a combination of in vitro and in silico techniques, we have identified interacting determinants relevant to the CaV3.3/genistein complex formation and propose that genistein directly blocks the function of the human CaV3.3 channel as a result of such interaction. Specifically, we proposed that a combination of polar interactions involving the three hydroxyl groups of genistein and an aromatic interaction with the fused rings are the main binding interactions in the complex formation. Our results pave the way for the rational development of improved and novel low-voltage activated T-type calcium channel inhibitors.

Attenuation of Native Hyperpolarization-Activated, Cyclic Nucleotide-Gated Channel Function by the Volatile Anesthetic Sevoflurane in Mouse Thalamocortical Relay Neurons

As thalamocortical relay neurons are ascribed a crucial role in signal propagation and information processing, they have attracted considerable attention as potential targets for anesthetic modulation. In this study, we analyzed the effects of different concentrations of sevoflurane on the excitability of thalamocortical relay neurons and hyperpolarization-activated, cyclic-nucleotide gated (HCN) channels, which play a decisive role in regulating membrane properties and rhythmic oscillatory activity. The effects of sevoflurane on single-cell excitability and native HCN channels were investigated in acutely prepared brain slices from adult wild-type mice with the whole-cell patch-clamp technique, using voltage-clamp and current-clamp protocols. Sevoflurane dose-dependently depressed membrane biophysics and HCN-mediated parameters of neuronal excitability. Respective half-maximal inhibitory and effective concentrations ranged between 0.30 (95% CI, 0.18–0.50) mM and 0.88 (95% CI, 0.40–2.20) mM. We witnessed a pronounced reduction of HCN dependent Ih current amplitude starting at a concentration of 0.45 mM [relative change at −133 mV; 0.45 mM sevoflurane: 0.85 (interquartile range, 0.79–0.92), n = 12, p = 0.011; 1.47 mM sevoflurane: 0.37 (interquartile range, 0.34–0.62), n = 5, p < 0.001] with a half-maximal inhibitory concentration of 0.88 (95% CI, 0.40–2.20) mM. In contrast, effects on voltage-dependent channel gating were modest with significant changes only occurring at 1.47 mM [absolute change of half-maximal activation potential; 1.47 mM: −7.2 (interquartile range, −10.3 to −5.8) mV, n = 5, p = 0.020]. In this study, we demonstrate that sevoflurane inhibits the excitability of thalamocortical relay neurons in a concentration-dependent manner within a clinically relevant range. Especially concerning its effects on native HCN channel function, our findings indicate substance-specific differences in comparison to other anesthetic agents. Considering the importance of HCN channels, the observed effects might mechanistically contribute to the hypnotic properties of sevoflurane.

Structure-Function of TMEM16 Ion Channels and Lipid Scramblases.

The TMEM16 protein family comprises two novel classes of structurally conserved but functionally distinct membrane transporters that function as Ca2+-dependent Cl- channels (CaCCs) or dual functional Ca2+-dependent ion channels and phospholipid scramblases. Extensive functional and structural studies have advanced our understanding of TMEM16 molecular mechanisms and physiological functions. TMEM16A and TMEM16B CaCCs control transepithelial fluid transport, smooth muscle contraction, and neuronal excitability, whereas TMEM16 phospholipid scramblases mediate the flip-flop of phospholipids across the membrane to allow phosphatidylserine externalization, which is essential in a plethora of important processes such as blood coagulation, bone development, and viral and cell fusion. In this chapter, we summarize the major methods in studying TMEM16 ion channels and scramblases and then focus on the current mechanistic understanding of TMEM16 Ca2+- and voltage-dependent channel gating as well as their ion and phospholipid permeation.

Inversion in a four-terminal superconducting device on the quartet line. I. Two-dimensional metal and the quartet beam splitter

In connection with the recent Harvard group experiment on graphene-based four-terminal Josephson junctions containing a grounded loop, we consider voltage biasing at opposite voltages on the quartet line and establish lowest-order perturbation theory in the tunnel amplitudes between a two-dimensional (2D) metal and four superconducting leads in the dirty limit. We present in addition general nonperturbative and nonadiabatic results. The critical current on the quartet line ${I}_{c}(\mathrm{\ensuremath{\Phi}}/{\mathrm{\ensuremath{\Phi}}}_{0})$ depends on the reduced flux $\mathrm{\ensuremath{\Phi}}/{\mathrm{\ensuremath{\Phi}}}_{0}$ via interference between the three-terminal quartets (3TQs) and the nonstandard four-terminal split quartets (4TSQs). The 4TSQs result from synchronizing two Josephson junctions by exchange of two quasiparticles ``surfing'' on the 2D quantum wake, and this mechanism is already operational at equilibrium. Perturbation theory in the tunnel amplitudes shows that the 3TQs are $\ensuremath{\pi}$-shifted but the 4TSQs are 0-shifted if the contacts have linear dimension which is large compared to the elastic mean free path. We establish the gate voltage dependence of the quartet critical current oscillations ${I}_{c}(\mathrm{\ensuremath{\Phi}}/{\mathrm{\ensuremath{\Phi}}}_{0})$. It is argued that ``observation of ${I}_{c}(0)\ensuremath{\ne}{I}_{c}(1/2)$'' implies ``evidence for the four-terminal 4TSQ'' for finite bias voltage on the quartet line and arbitrary interface transparencies. This statement relies on physically motivated approximations leading to the Ambegaokar-Baratoff-type formula for the quartet critical current-flux relation. It is concluded that the recent experiment mentioned above finds evidence for the four-terminal 4TSQ.

Disrupting polycystin-2 EF hand Ca2+ affinity does not alter channel function or contribute to polycystic kidney disease.

ABSTRACTApproximately 15% of autosomal dominant polycystic kidney disease (ADPKD) is caused by variants in PKD2. PKD2 encodes polycystin-2, which forms an ion channel in primary cilia and endoplasmic reticulum (ER) membranes of renal collecting duct cells. Elevated internal Ca2+ modulates polycystin-2 voltage-dependent gating and subsequent desensitization – two biophysical regulatory mechanisms that control its function at physiological membrane potentials. Here, we refute the hypothesis that Ca2+ occupancy of the polycystin-2 intracellular EF hand is responsible for these forms of channel regulation, and, if disrupted, results in ADPKD. We identify and introduce mutations that attenuate Ca2+-EF hand affinity but find channel function is unaltered in the primary cilia and ER membranes. We generated two new mouse strains that harbor distinct mutations that abolish Ca2+-EF hand association but do not result in a PKD phenotype. Our findings suggest that additional Ca2+-binding sites within polycystin-2 or Ca2+-dependent modifiers are responsible for regulating channel activity.

Sensitive detection of water/oxygen molecule adsorption and reaction on a titanium oxide nanosheet with a graphene field effect transistor

We have investigated molecule adsorption phenomena on a chemically active surface of titanium oxide nanosheet by coupling with an electrically sensitive graphene field effect transistor (FET). Super-hydrophilic surface of the titanium oxide nanosheet forms a water-layer in ambient air which exhibits a large hysteresis of drain current in the hybrid FET for sweeping gate-voltage. The large hysteresis disappears in vacuum, which indicates physically adsorbed water molecules on the surface of the titanium oxide nanosheet dominantly contribute to the hysteresis. UV light irradiation in vacuum significantly changes the drain current due to desorption of the adsorbed molecules. Sufficient UV irradiation results in symmetric gate-voltage dependence similar to those of conventional graphene FETs. Exposure to an oxygen gas atmosphere leads to a heavy hole doping in the FET, where the binding of the oxygen molecules is stronger than that of water molecules. In a humidified nitrogen atmosphere, a large shift of charge neutrality point is observed in transfer characteristics crossing between electron doping and hole doping. By contrast, a clear square-shaped hysteresis loop is observed in a humidified oxygen atmosphere, where the hole density in the graphene drastically changed with O2/H2O redox couple reaction on the titanium oxide nanosheet.