Peter Pohl

• MD
• Professor at Johannes Kepler University of Linz

Native gel electrophoresis techniques, such as blue or clear native gel electrophoresis (BNE or CNE), are widely used to separate and characterize proteins. However, in high-resolution CNE, mild anionic or neutral detergents are often used at concentrations too low to prevent membrane-protein aggregation. Additionally, the identification of protein...

Fluorescent labeling of membrane proteins is essential for exploring their functions, signaling pathways, interaction partners, and structural dynamics. Organic fluorophores are commonly used for this purpose due to their favorable photophysical properties and photostability. However, a persistent challenge is the inaccessibility of the surface-exp...

Voltage-gated potassium (Kv) channels shape action potentials, with their activity dependent on anionic lipids and modulated by membrane tension. However, the mechanism by which lipids regulate channel gating is not entirely clear. Cryo-electron microscopy studies suggest that in the down state— typically associated with the inactive channel—argini...

Controlling the flux of specific ions across lipid bilayers is a central challenge for cellular life. Here, we present a general strategy for the precise, noninvasive, and reversible photoregulation of specific ion transport across lipid bilayer membranes: using embedded azobenzene-containing photolipids as dopants that effectively photo-control th...

Proton translocation through lipid membranes is a fundamental process in the field of biology. Several theoretical models have been developed and presented over the years to explain the phenomenon, yet the exact mechanism is still not well understood. Here, we show that proton translocation is directly related to membrane potential fluctuations. Us...

Optically-induced changes in membrane capacitance may regulate neuronal activity without requiring genetic modifications. Previously, they mainly relied on sudden temperature jumps due to light absorption by membrane-associated nanomaterials or water. Yet, nanomaterial targeting or the required high infrared light intensities obstruct broad applica...

The universally conserved protein YidC aids in the insertion and folding of transmembrane polypeptides. Supposedly, a charged arginine faces its hydrophobic lipid core, facilitating polypeptide sliding along YidC’s surface. How the membrane barrier to other molecules may be maintained is unclear. Here, we show that the purified and reconstituted E....

The high propensity of protons to stay at interfaces has attracted much attention over the decades. It enables long-range interfacial proton diffusion without relying on titratable residues or electrostatic attraction. As a result, various phenomena manifest themselves, ranging from spillover in material sciences to local proton circuits between pr...

Optically-induced changes in membrane capacitance may regulate neuronal activity without requiring genetic modifications. Previously, they mainly relied on sudden temperature jumps due to light absorption by membrane-associated nanomaterials or water. Yet, nanomaterial targeting or the required high infrared light intensities obstruct broad applica...

The universally conserved protein YidC aids the insertion and folding of transmembrane polypeptides independently or as a part of the SecYEG translocon complex. In the former scenario, the lipid-exposed YidC surface equipped with a highly conserved positively charged arginine is thought to facilitate membrane insertion of the nascent chain by provi...

Millions of years of evolution have produced proteinaceous water channels (aquaporins) that combine perfect selectivity with a transport rate at the edge of the diffusion limit. However, Itoh et al. recently claimed in Science that artificial channels are 100 times faster and almost as selective. The published deflation kinetics of vesicles contain...

Proton relay between interfacial water molecules allows rapid two-dimensional diffusion. An energy barrier, ΔGr‡, opposes proton-surface-to-bulk release. The ΔGr‡-regulating mechanism thus far has remained unknown. Here, we explored the effect interfacial charges have on ΔGr‡’s enthalpic and entropic constituents, ΔGH‡ and ΔGS‡, respectively. A lig...

The voltage‐gated proton channel, HV1, 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...

In concert with irreversible non-equilibrium peptide translation by the ribosome, the nascent polypeptide chain may integrate into the membrane or translocate to the other side of the membrane, facilitated by the conserved protein translocation channel SecYEG in bacteria and Sec61 in eukaryotes. Assuming equilibrium for the decision processes yield...

Yeast S. cerevisiae has been shown to suppress a sterol biosynthesis as a response to hyperosmotic stress. In the case of sodium stress, the failure to suppress biosynthesis leads to an increase in cytosolic sodium. The major yeast sterol, ergosterol, is known to regulate functioning of plasma membrane proteins. Therefore, it has been suggested tha...

The chase toward endowing chemical compounds with machine-like functions mimicking those of biological molecular machineries has yielded a variety of artificial molecular motors (AMMs). Pharmaceutical applications of photoexcited monomolecular unidirectionally-rotating AMMs have been envisioned in view of their ability to permeabilize biological me...

16 17 The voltage-gated proton channel, HV1, is crucial for innate immune responses. According to 18 alternative hypotheses, protons either hop on top of an uninterrupted water wire or bypass titratable 19 amino acids, interrupting the water wire halfway across the membrane. To distinguish between both 20 hypotheses, we estimate the water mobility...

Lipid domains less than 200 nm in size may form a scaffold, enabling the concerted function of plasma membrane proteins. The size-regulating mechanism is under debate. We tested the hypotheses that large values of spontaneous monolayer curvature are incompatible with micrometer-sized domains. Here, we used the transition of photoswitchable lipids f...

Various nanoscopic channels of roughly equal diameter and length facilitate single-file diffusion at vastly different rates. The underlying variance of the energetic barriers to transport is poorly understood. First, water partitioning into channels so narrow that individual molecules cannot overtake each other incurs an energetic penalty. Correspo...

It is my pleasure to write a few words to introduce myself to the readers of Biophysical Reviews as part of the “Meet the Councilor Series.” Currently, I am serving the second period as IUPAB councilor after having been elected first in 2017. Initially, I studied Biophysics in Moscow (Russia) and later Medicine in Halle (Germany). My scientific car...

Membrane proteins can be regulated by alterations in material properties intrinsic to the hosting lipid bilayer. Here, we investigated whether the reversible photoisomerization of bilayer-embedded diacylglycerols (OptoDArG) with two azobenzene-containing acyl chains may trigger such regulatory events. We observed an augmented open probability of th...

Many membrane channels and transporters conduct water. Their reconstitution into giant unilamellar vesicles (GUVs) – here represented by the water tap – serves to assess their unitary hydraulic permeabilities. Boytsov, Hannesschlaeger, Horner, Siligan, and Pohl report the assay in article 1900450. It is based on measurements of (i) GUV volume chang...

Translocation of many secretory proteins through the bacterial plasma membrane is facilitated by a complex of the SecYEG channel with the motor protein SecA. The ATP-free complex is unstable in detergent, raising the question how SecA may perform several rounds of ATP hydrolysis without being released from the membrane embedded SecYEG. Here we show...

Archaeal single-cell microorganisms are characterized by particularly strong resistance to harsh environments at extreme temperature ranges, pH values, high pressure and salinity. The recognition of molecules rendering microorganisms’ membranes stable in such conditions represents an important step towards understanding of the molecular mechanisms...

Measurements of the unitary hydraulic conductivity of membrane channels, pf, may be hampered by difficulties in producing sufficient quantities of purified and reconstituted proteins. Low yield expression, the purely empiric choice of detergents, as well as protein aggregation and misfolding during reconstitution may result in an average of less th...

Lipid rafts serve as anchoring platforms for membrane proteins. Thus far they escaped direct observation by light microscopy due to their small size. Here we used differently colored dyes as reporters for the registration of both ordered and disordered lipids from the two leaves of a freestanding bilayer. Photoswitchable lipids dissolved or reforme...

Ordered lipid domains (OLD) may serve as signaling platforms provided both membrane leaflets contribute. Interleaflet registration of phase separated regions in symmetric bilayers is driven by (i) the line tension around the thicker ordered domains (1) and (ii) membrane undulations (2, 3). How these forces act to generate membrane spanning domains...

The bacterial channel SecYEG efficiently translocates both hydrophobic and hydrophilic proteins across the plasma membrane. Translocating polypeptide chains may dislodge the plug, a half helix that blocks the permeation of small molecules, from its position in the middle of the aqueous translocation channel. Instead of the plug, six isoleucines in...

Translocation of many secretory proteins through the bacterial plasma membrane is facilitated by the protein translocation complex SecYEG. The motor protein SecA is thought to use ATP hydrolysis to push the polypeptide chain through SecYEG. How SecA may perform several rounds of ATP hydrolysis is unclear, since its nucleotide-free conformation was...

Spontaneous solute and solvent permeation through membranes is of vital importance to human life, be it gas exchange in red blood cells, metabolite excretion, drug/ toxin uptake, or water homeostasis. Knowledge of the underlying molecular mechanisms is the sine qua non of every functional assignment to membrane transporters. The basis of our curren...

The assessment of weak acid membrane permeability (Pm) frequently involves large unilamellar vesicles. It relies on measurements of the intravesicular pH drop, ΔpHin, in response to a sudden augmentation of external acid concentration. However, ΔpHin may be primarily governed by non-instantaneous protonation and deprotonation reactions of (i) the a...

This is a perspective article entitled “Frontiers in computational biophysics: understanding conformational dynamics of complex lipid mixtures relevant to biology” which is following a CECAM meeting with the same name. Graphical Abstract

The membrane permeability P of organic ions was reported to be governed by the structure of the permeating molecule. Thus far, it is unclear whether the ion structure alters membrane partition or translocation proper across the membrane. Here, we obtained P values for 24 anionic compounds (18 concrete values, 6 upper limits) measuring the current t...

Photodynamic therapy uses photosensitizers (PS) to kill cancer cells by generating reactive oxygen species - like singlet oxygen (SO) - upon illumination with visible light. PS membrane anchoring augments local SO concentration, which in turn increases photodynamic efficiency. The latter may suffer from SO's escape into the aqueous solution or prem...

Vitamin C (VC)—a collective term for the different oxidation and protonation forms of ascorbic acid (AscH)—is an essential micronutrient that serves as (i) a potent antioxidant and (ii) a cofactor of a manifold of enzymatic processes. Its role in health is related to redox balance maintenance, which is altered in diseases such as obesity, cancer, n...

Water at interfaces governs many processes from the molecular scale of electrochemical and enzymatic reactions to protein folding. Here we focus on water transport through proteinaceous pores that are so narrow that the water molecules cannot overtake each other in the pore. After a short introduction into the single-file transport theory, we analy...

This review focusses on the energetics of protein translocation via the Sec translocation machinery. First we complement structural data about SecYEG’s conformational rearrangements by insight obtained from functional assays. These include measurements of SecYEG permeability that allow assessment of channel gating by ligand binding and membrane vol...

Long-chain free fatty acids (FFAs) play an important role in several physiological and pathological processes such as lipid fusion, adjustments of membrane permeability and fluidity, and the regulation of enzyme and protein activities. FFA-facilitated membrane proton transport (flip-flop) and FFA-dependent proton transport by membrane proteins (e.g...

Water molecules lose two of their four bulk neighbours when entering single-file channels. This process may be sensitive to the presence of positive and negative charges at the channel mouth, since the costs for dehydrating cations and anions differ by a large margin. However, it is not known whether entrance charges affect single channel water per...

Significance Lipid bilayer membranes are responsible for compartmentalization, signaling, transport, and flow of charge in living cells. Membranes self-assemble in aqueous solutions. Without a hydrating environment, membranes cannot exist. It is therefore surprising to note that the hydrating water is neglected in most membrane-related studies. We...

Tunuguntla et al. (Reports, 25 August 2017, p. 792) report that permeation of single-file water occurs faster through carbon nanotubes than through aquaporins. We show that this conclusion violates fundamental thermodynamic laws: Because of its much lower activation energy, aquaporin-mediated water transport must be orders of magnitude faster. Leak...

Proton transport at water/membrane interfaces plays a fundamental role for a myriad of bioenergetic processes. Here we have performed ab initio molecular dynamics simulations of proton transfer along two phosphatidylcholine bilayers. As found in previous theoretical studies, the excess proton is preferably located at the water/membrane interface. F...

Supporting text, figures and tables. (PDF)

The heterotrimeric SecYEG complex cooperates with YidC to facilitate membrane protein insertion by an unknown mechanism. Here we show that YidC contacts the interior of the SecY channel resulting in a ligand-activated and voltage-dependent complex with distinct ion channel characteristics. The SecYEG pore diameter decreases from 8 Å to only 5 Å for...

Proton diffusion along biological membranes is vitally important for cellular energetics. Here we extended previous time-resolved fluorescence measurements to study the time and temperature dependence of surface proton transport. We determined the Gibbs activation energy barrier ΔG‡r that opposes proton surface-to-bulk release from Arrhenius plots...

The mechanism of action of photosensitizers used in the photodynamic tumor therapy is studied by using bilayer lipid membranes (BLM) which adequately simulate the plasma membranes of cells. The formation of singlet oxygen upon photoexcitation of photosensitizer (tetrasulfonated alumophthalocyanine) is revealed by the photosensitizer-induced decompo...

The small intestine is void of aquaporins adept at facilitating vectorial water transport, and yet it reabsorbs ∼8 liters of fluid daily. Implications of the sodium glucose cotransporter SGLT1 in either pumping water or passively channeling water contrast with its reported water transporting capacity, which lags behind that of aquaporin-1 by 3 orde...

Phase separation in biological membranes plays an important role in protein targeting and transmembrane signaling. Its occurrence in both membrane leaflets commonly gives rise to matching liquid or liquid-ordered domains in the opposing monolayers. The underlying mechanism of such co-localization is not fully understood. The decrease of the line te...

High-speed atomic force microscopy (HS-AFM) can be used to visualize function-related conformational changes of single soluble proteins. Similar studies of single membrane proteins are, however, hampered by a lack of suitable flat, non-interacting membrane supports and by high protein mobility. Here we show that streptavidin crystals grown on mica-...

Understanding the structure and dynamics of water's constituent ions, proton and hydroxide, has been a subject of numerous experimental and theoretical studies over the last century. Besides their obvious importance in acid-base chemistry, these ions play an important role in numerous applications ranging from enzyme catalysis to environmental chem...

Understanding the structure and dynamics of water’s constituent ions, proton and hydroxide, has been a subject of numerous experimental and theoretical studies over the last century. Besides their obvious importance in acid−base chemistry, these ions play an important role in numerous applications ranging from enzyme catalysis to environmental chem...

Water is an essential participant in the stability, structure, dynamics, and function of proteins and other biomolecules. Thermodynamically, changes in the aqueous environment affect the stability of biomolecules. Structurally, water participates chemically in the catalytic function of proteins and nucleic acids and physically in the collapse of th...

The small intestine is void of aquaporins adept at facilitating vectorial water transport and yet it reabsorbs about eight liters of fluid daily. Implications of the sodium glucose cotransporter SGLT1 in either pumping water or passively channeling water, contrast with its reported water transporting capacity, which lags behind that of aquaporin-1...

DOI:http://dx.doi.org/10.1103/PhysRevLett.116.079802

The universally conserved protein YidC functions as a membrane protein insertase. It facilitates transmembrane domains’ passage out of SecYEG's lateral gate and inserts short or closely spaced membrane proteins independent of SecYEG. According to a recent cryo-EM density map of a ribosome-nascent chain complex (RNC) bound to YidC in detergent in ad...

The small intestine is void of aquaporins adept at facilitating vectorial water transport and yet it reabsorbs about eight liters of fluid daily. Implications of the sodium glucose cotransporter SGLT1 in either pumping water or passively channeling water, contrast with its reported water transporting capacity, which lags behind that of aquaporin-1...

The mechanism responsible for the registration of liquid-ordered (Lo) domains in the two membrane leaflets is a matter of debate. As an alternative to the thus far enigmatic interactions at the membrane midplane, we propose that minimization of the line tension around the thicker Lo bilayer drives registration. Based on the continuum elasticity the...

The heterotrimeric protein translocation channel SecYEG enables (i) soluble proteins to cross the inner membrane and (ii) hydrophobic proteins to enter the membrane interior. It contains an aqueous pore that, in its resting state, is sealed by a ring of six hydrophobic residues and a half helix, termed the plug [1]. Signal sequence binding or ribos...

Aquaporins are conserved throughout all kingdoms of life. Each aquaporin monomer possesses a channel that facilitates the transport of water across the membrane, and yet the protein assembles into a tetramer. We investigated the physiological function of the tetrameric assembly by introducing destabilizing point mutations. Therefore, we overexpress...