Trapped Pore Waters in the Open Proton Channel H_V1

Abstract

to H V 1 gating. [3] Its structure is homologous to classical voltage-gated channels' voltage-sensing domain (VSD). [4] Experimentally, exclusively closed structures are available -a crystal structure of a chimera between mouse H V 1 and Ciona intestinalis voltage-sensing phosphatase [5] and an NMR structure of the human H V 1. [6] Also, electron paramagnetic resonance (EPR) spectroscopy data decipher the resting structure of H V 1. [7] An AlphaFold structure has been recently added (Figure 1). AlphaFold uses a machine-learning approach. [8] Its deep learning algorithm exploits physical and biological knowledge about protein structure -mainly gained by X-ray crystallography, cryo-electron microscopy, or NMR. None of these technical approaches allows the application of membrane voltage, a prerequisite to attain the open channel configuration of voltage-sensitive channels. Since Hv1 belongs to this class of channels, the AlphaFold structure reflects, in all likelihood, the closed channel structure.The analysis of the AlphaFold structure corroborates the conclusion (Figure 1A,B). It shows three positively charged arginine residues in the membrane-spanning part of the 4th helix (S4) responsible for Hv1's voltage sensitivity. They may interact with three negatively charged aspartate residues on the first (S1) and third helices (S3). The AlphaFold structure suggests the following Coulombic interactions: R205 -D112 (on S1), R211 -D185 (on S3) and R208 -D174 (on S3). There is consensus that i) D174 is part of the intracellular electrostatic network [9,10] and ii) D174's salt bridge with R208 stabilizes the closed, resting-state conformation. [11] The open channel's structure has solely been computed as homology models. [12][13][14][15][16][17][18] One of the most recent models reflects the experimentally estimated gating charge suggesting that the transition from the closed to the open conformation may be correctly captured. [17] Yet, the closed conformations of the homology model (Figure S1, Supporting Information) and AlphaFold (Figure 1A,B) differ in pore diameter (Figure 1C). While the former is large enough to allow water molecules to pass, the latter is much too narrow. The observation suggests that an assessment of H V 1's unitary water permeability, p f , may be used as a diagnostic tool. By undertaking water flux measurements through purified and reconstituted H V 1 channels in The voltage-gated proton channel, H V 1, is crucial for innate immune responses. According to alternative hypotheses, protons either hop on top of an uninterrupted water wire or bypass titratable amino acids, interrupting the water wire halfway across the membrane. To distinguish between both hypotheses, the water mobility for the putative case of an uninterrupted wire is estimated. The predicted single-channel water permeability 2.3 × 10 −12 cm 3 s −1 reflects the permeability-governing number of hydrogen bonds between water molecules in single-file configuration and pore residues. However, the measured unitary water permeability does no...

Proton Paths in Models of the Hv1 Proton Channel

“…The proposed closed structure of Geragotelis et al is also too permeable. The same conclusion was reached by looking at the pore dimensions . Perhaps the voltage that was applied in the MD simulation caused a transient opening of the structure, which then did not have enough time to relax.…”

“…The same conclusion was reached by looking at the pore dimensions. 53 Perhaps the voltage that was applied in the MD simulation caused a transient opening of the structure, which then did not have enough time to relax. Concerning the open state models, we can safely say that the one R3D model (gerO) is much more easily permeable than the several R2D models we have considered.…”

“…Another argument in favor of the shuttling hypothesis was that no water permeability is observed. 53 However, it is not clear whether water-wire transport must always be associated with water movement. This issue needs to be further investigated in the future.…”

Voltage-Gated Proton Channels in the Tree of Life

“…Multiple evidence as the high temperature dependence of H V [47,87], the diminished deuterium conductance [83], the H + hopping between pairs of charged residues lining H V pore detected in QM/MM molecular dynamics simulations [152], or more recently the proposed existence of an interrupted water wire within H V 1 pore [153], suggest that a protonable carboxyl group is essential for the selectivity of H V channels. However, the mechanism underlying this selectivity involves also specific physicochemical microenvironments where H + conduction and selectivity are coupled [120,132,141,[154][155][156][157][158].…”

Proton channels in molluscs: A new bivalvian‐specific minimal HV4 channel

“…pK a values of internal lysine residues in nucleases reported to be as low as 5.3 depending on the polarity of its microenvironment [50]. Studies conducted on proton channels have demonstrated that the arginine residue located at the third position (R3) of the voltagesensing motif (VSM) is exposed to the cytosolic solution when the channel is in its resting state [42,49,[51][52][53][54][55]. Under conditions of negligible pK a changes due to microenvironmental effects of the CgH V 4 structure on K3, our experimental settings permit protonation of the terminal amino group of Lys 150 , which would act as a discrete gating charge [56].…”