Recent X-ray crystal structures of the two-pore domain (K2P) family of potassium channels have revealed a unique structural architecture at the point where the cytoplasmic bundle-crossing gate is found in most other tetrameric K+ channels. However, despite the apparently open nature of the inner pore in the TWIK-1 (K2P1/KCNK1) crystal structure, the reasons underlying its low levels of functional activity remain unclear. In this study, we use a combination of molecular dynamics simulations and functional validation to demonstrate that TWIK-1 possesses a hydrophobic barrier deep within the inner pore, and that stochastic dewetting of this hydrophobic constriction acts as a major barrier to ion conduction. These results not only provide an important insight into the mechanisms which control TWIK-1 channel activity, but also have important implications for our understanding of how ion permeation may be controlled in similar ion channels and pores.

Performance of a sensing device is dependent on its construction material, especially for components that are directly involved in transporting and translating signals across the device. Understanding the morphology and characteristics of the material components is therefore crucial in the development of any sensing device. This work examines the morphological and electrochemical characteristics of reduced graphene oxide interspersed with poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (rGO-PEDOT:PSS) used as a transducer material deposited on a commercially available screen-printed carbon electrode (SPCE). Electron microscopy shows that PEDOT:PSS is interspersed between rGO layers. Raman and XRD analyses suggest that the graphene crystallinity in GO-PEDOT:PSS and rGO-PEDOT:PSS remains intact. Instead, PEDOT:PSS undergoes a change in structure to allow PEDOT to blend into the graphene structure and partake in the π-π interaction with the surface of the rGO layers. Incorporation of PEDOT:PSS also appears to improve the electrochemical behavior of the composite, leading to a higher peak current of 1.184 mA, as measured by cyclic voltammetry, compared to 0.522 mA when rGO is used alone. The rGO-PEDOT:PSS transducing material blended with glucose oxidase was tested for glucose detection. The sensitivity of glucose detection was shown to be 57.3 µA/(mM·cm2) with a detection limit of 86.8 µM.

Several recent ion channel structures have revealed large side portals, or ‘fenestrations’ at the interface between their transmembrane helices that potentially expose the ion conduction pathway to the lipid core of the bilayer. In a recent study we demonstrated that functional activity of the TWIK-1 K2P channel is influenced by the presence of hydrophobic residues deep within the inner pore. These residues are located near the fenestrations in the TWIK-1 structure and promote dewetting of the pore by forming a hydrophobic barrier to ion conduction. During our previous MD simulations, lipid tails were observed to enter these fenestrations. In this addendum to that study, we investigate lipid contribution to the dewetting process. Our results demonstrate that lipid tails from both the upper and lower leaflets can occupy the fenestrations and partially penetrate into the pore. The lipid tails do not sterically occlude the pore, but there is an inverse correlation between the presence of water within the hydrophobic barrier and the number of lipids tails within the lining of the pore. However, dewetting still occurs in the absence of lipids tails, and pore hydration appears to be determined primarily by those side-chains lining the narrowest part of the pore cavity.

TASK-2, a member of the Two-Pore Domain (K2P) subfamily of K+ channels, is encoded by the KCNK5 gene. The channel is expressed primarily in renal epithelial tissues and a potentially deleterious missense variant in KCNK5 has recently been shown to be prevalent amongst patients predisposed to the development of Balkan Endemic Nephropathy (BEN), a chronic tubulointerstitial renal disease of unknown etiology. In this study we show that this variant (T108P) results in a complete loss of channel function and is associated with a major reduction in TASK-2 channel subunits at the cell surface. Furthermore, these mutant subunits have a suppressive or ‘dominant-negative’ effect on channel function when coexpressed with wild-type subunits. This missense variant is located at the extracellular surface of the M2 transmembrane helix and by using a combination of structural modelling and further functional analysis we also show that this highly-conserved threonine residue is critical for the correct function of other K2P channels. These results therefore provide further structural and functional insights into the possible pathophysiological effects of this missense variant in TASK-2.

Electron-transfer kinetics and impedance at the electrode-solution interface affect biosensor performance. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) are used to understand the reversibility of electron transfer and impedance at the electrode-solution interface, respectively. Effective surface areas calculated based on the Randles-Sevcik equation for a bare screen-printed carbon electrode (SPCE), a graphene oxide (GO)-poly (3,4-ethylenedioxythiophene):polystyrenesulfonic acid (PEDOT:PSS)-modified electrode (GO-PEDOT:PSS/SPCE), a Engineering main heading: Conducting polymers

The present work describes a self-sustaining bioelectrochemical system that adopts simple cell configurations and operates in uncontrolled ambient surroundings. The microbial fuel cell (MFC) was comprised of white-rot fungus of Phanaerochaete chrysosporium fed with oil palm empty fruit bunch (EFB) as the substrate. This fungal strain degrades lignin by producing ligninolytic enzymes such as laccase, which demonstrates a specific affinity for oxygen as its electron acceptor. By simply pairing zinc and the air electrode in a membraneless, single-chamber, 250-mL enclosure, electricity could be harvested. The microbial zinc/air cell is capable of sustaining a 1 mA discharge current continuously for 44 days (i.e., discharge capacity of 1056 mAh). The role of the metabolic activities of P. chrysosporium on EFB towards the MFC’s performance is supported by linear sweep voltammetry measurement and scanning electron microscopy observations. The ability of the MFC to sustain its discharge for a prolonged duration despite the fungal microbes not being attached to the air electrode is attributed to the formation of a network of filamentous hyphae under the submerged culture. Further, gradual lignin decomposition by fungal inocula ensures a continuous supply of laccase enzyme and radical oxidants to the MFC. These factors promote a self-sustaining MFC devoid of any control features.

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Cyclic voltammetry (CV) and electrical impedance spectroscopy (EIS) are electrochemical techniques to characterize reversibility of electron transfer and impedance at the electrode-solution interface, respectively. Reduced graphene oxide (rGO) and conductive polymer PEDOT:PSS are often used to enhance electron transfer at electrode surface. This chapter provides a step-by-step methodology of CV and EIS conducted on screenprinted carbon electrode (SPCE) modified with rGO-PEDOT:PSS and brief discussion on the CV and EIS results. The CV of rGO-PEDOT:PSS shows a reversible electron transfer in comparison to SPCE modified with PEDOT:PSS. For EIS, rGO-PEDOT:PSS was found to reduce the Warburg impedance, suggesting enhanced electrode conductivity. These results suggest that rGO-PEDOT:PSS is a suitable material for biosensing purpose.