Molecular basis of SARS-CoV-2 Omicron variant receptor engagement and antibody evasion and neutralization

The SARS-CoV-2 Omicron variant exhibits striking immune evasion and is spreading globally at an unprecedented speed. Understanding the underlying structural basis of the high transmissibility and greatly enhanced immune evasion of Omicron is of high importance. Here through cryo-EM analysis, we present both the closed and open states of the Omicron spike, which appear more compact than the counterparts of the G614 strain, potentially related to the Omicron substitution induced enhanced protomer-protomer and S1-S2 interactions. The closed state showing dominant population may indicate a conformational masking mechanism of immune evasion for Omicron spike. Moreover, we capture two states for the Omicron S/ACE2 complex with S binding one or two ACE2s, revealing that the substitutions on the Omicron RBM result in new salt bridges/H-bonds and more favorable electrostatic surface properties, together strengthened interaction with ACE2, in line with the higher ACE2 affinity of the Omicron relative to the G614 strain. Furthermore, we determine cryo-EM structures of the Omicron S/S3H3 Fab, an antibody able to cross-neutralize major variants of concern including Omicron, elucidating the structural basis for S3H3-mediated broad-spectrum neutralization. Our findings shed new lights on the high transmissibility and immune evasion of the Omicron variant and may also inform design of broadly effective vaccines against emerging variants.

Structural insights into the mechanism of nucleotide regulation of pancreatic KATP channel

ATP-sensitive potassium channels (KATP) are metabolic sensors that convert the intracellular ATP/ADP ratio to the excitability of cells. They are involved in many physiological processes and implicated in several human diseases. Here we present the cryo-EM structures of the pancreatic KATP channel in both the closed state and the pre-open state, resolved in the same sample. The nucleotides bind at the inhibitory sites of the Kir6.2 channel in the closed state but not in the pre-open state. Structural comparisons reveal the mechanism for ATP inhibition and Mg-ADP activation, two fundamental properties of KATP channels. Moreover, the structure also uncovers the activation mechanism of diazoxide-type KATP openers.

Piezoelectric Normally Open Microvalve with Multiple Valve Seat Trenches for Medical Applications

Microfluidic systems for medical applications necessitate reliable, wide flow range, and low leakage microvalves for flow path control. High design complexity of microvalves increases the risk of possible malfunction. We present a normally open microvalve based on energy-efficient piezoelectric actuation for high closing forces and micromachined valve seat trenches for reliable valve operation. A comprehensive investigation of influencing parameters is performed by extensive fluidic 3D finite element simulation, derivation of an analytical closed state leakage rate model, as well as fabrication and test of the microvalve. Additional valve seat coating and a high force actuator are introduced for further leakage reduction. The microvalve has a wide-open flow range as well as good sealing abilities in closed state. Extensive fatigue tests of 1 × 106 actuation cycles show that additional coating of the valve seat or increased actuator strength promote sealing performance stability. Analytical calculations of leakage are suitable to estimate experimentally obtained leakage rates and, along with computational fluidic dynamic (CFD) simulations, enable future microvalve design optimization. In conclusion, we demonstrate that the presented normally open microvalve is suitable for the design of safe and reliable microfluidic devices for medical applications.

Cryo-EM structure of the human TACAN channel in a closed state

TACAN is an ion channel involved in sensing mechanical pain. It has recently been shown to represent a novel and evolutionarily conserved class of mechanosensitive channels. Here, we present the cryoelectron microscopic structure of human TACAN (hTACAN). hTACAN forms a dimer in which each protomer consists of a transmembrane globular domain (TMD) that is formed of six helices and an intracellular domain (ICD) that is formed of two helices. Molecular dynamic simulations suggest that a putative ion conduction pathway is located inside each protomer. Single point mutation of the key residue Met207 significantly increased the surface tension activated currents. Moreover, cholesterols were identified at the flank of each subunit. Our data show the molecular assembly of hTACAN and suggest that the wild type hTACAN is in a closed state, providing a basis for further understanding the activation mechanism of the hTACAN channel.

Plasticity in AAA+ proteases reveals ATP-dependent substrate specificity principles

SummaryIn bacteria, AAA+ proteases such as Lon and ClpXP specifically degrade substrates to promote growth and stress responses. Here, we find that an ATP-binding mutant of ClpX suppresses physiological defects of a Lon-deficient strain by shifting ClpXP protease specificity toward normally Lon-restricted substrates and away from normal ClpXP targets. Reconstitution with purified proteins assigns these effects to changes in direct recognition and processing of substrates. We show that wildtype ClpXP specificity can be similarly altered when ATP hydrolysis is reduced, which unexpectedly accelerates degradation of some substrates. This activation corresponds with changes in ClpX conformation, leading to a model where ClpX cycles between ‘capture’ and ‘processive’ states depending on ATP loading. Limiting ATP binding alters dynamics between states affording better recognition of unorthodox substrates, but worse degradation of proteins specifically bound by the processive state. Thus, AAA+ protease specificities can be directly tuned by differences in ATP hydrolysis rates.HighlightsA mutation in the Walker B region of ClpX induces recognition of new substrates.Proteases are optimized for specific functions but barrier to recognize new substrates is easily overcome.Expanding substrate recognition by a protease comes at the cost of reducing native substrate degradation.Decreasing ATP enhances ClpXP mediated degradation of certain classes of substrates.ClpX adopts distinct conformational states to favor better recognition of some substrates over others.Graphical AbstractIn a wildtype cell, AAA+ proteases Lon and ClpXP promote normal growth by degrading distinct substrates. ClpX*P can compensate for the absence of the Lon protease by tuning ClpXP substrate specificity to better degrade Lon-privileged substrates (such as DnaA, SciP, and misfolded proteins) but this comes at the cost of native ClpXP substrates (such as ssrA-tagged proteins and CtrA). We propose that ClpX alternates between a closed and open conformation and promoting one state over the other leads to alterations in substrate specificity. In the presence of ClpX* or in ATP-limited conditions, the open state is favored, allowing capture and recognition of substrates such as casein. The balance shifts to the closed state under high ATP conditions, allowing degradation of substrates such as GFP-ssrA, which preferentially bind the closed state.

Mathematical Modeling Reveals Quantitative Properties of KEAP1-NRF2 Signaling

In response to oxidative and electrophilic stresses, cells launch an NRF2-mediated transcriptional antioxidant program. The activation of NRF2 depends on a redox sensor, KEAP1, which acts as an E3-ligase adaptor to promote the ubiquitination and degradation of NRF2. While a great deal has been learned about the molecular details of KEAP1, NRF2, and their interactions, the quantitative aspects of signal transfer conveyed by this redox duo are still largely unexplored. In the present study, we examined the signaling properties including response time, half-life, maximal activation, and response steepness (ultrasensitivity) of NRF2, through a suite of mathematical models. The models describe, with increasing complexity, the reversible binding of KEAP1 dimer and NRF2 via the ETGE and DLG motifs, NRF2 production, KEAP1-dependent and independent NRF2 degradation, and perturbations by different classes of NRF2 activators. Our simulations revealed that at the basal condition, NRF2 molecules are largely sequestered by KEAP1, with the KEAP1-NRF2 complex comparably distributed in either an ETGE-bound only (open) state or an ETGE and DLG dual-bound (closed) state, corresponding to the unlatched and latched configurations of the conceptual hinge-latch model. With two-step ETGE binding, the open and closed states operate in cycle mode at the basal condition and transition to equilibrium mode at stressed conditions. Class I-V, electrophilic NRF2 activators, which modify redox-sensing cysteine residues of KEAP1, shift the balance to a closed state that is unable to degrade NRF2 effectively. Total NRF2 has to accumulate to a level that nearly saturates existing KEAP1 to make sufficient free NRF2, therefore introducing a signaling delay. At the juncture of KEAP1 saturation, ultrasensitive NRF2 activation, i.e., a steep rise in the free NRF2 level, can occur through two simultaneous mechanisms, zero-order degradation mediated by DLG binding and protein sequestration (molecular titration) mediated by ETGE binding. These response characteristics of class I-V activators do not require disruption of DLG binding to unlatch the KEAP1-NRF2 complex. In comparison, class VI NRF2 activators, which directly compete with NRF2 for KEAP1 binding, can unlatch or even unhinge the KEAP1-NRF2 complex. This causes a shift to the open state of KEAP1-NRF2 complex and ultimately its complete dissociation, resulting in a fast release of free NRF2 followed by stabilization. Although class VI activators may induce free NRF2 to higher levels, ultrasensitivity is lost due to lower free KEAP1 and thus its NRF2-sequestering effect. Stress-induced NRF2 nuclear accumulation is enhanced when basal nuclear NRF2 turnover constitutes a small load to NRF2 production. Our simulation further demonstrated that optimal abundances of cytosolic and nuclear KEAP1 exist to maximize ultrasensitivity. In summary, by simulating the dual role of KEAP1 in repressing NRF2, i.e., sequestration and promoting degradation, our mathematical modeling provides key novel quantitative insights into the signaling properties of the crucial KEAP1-NRF2 module of the cellular antioxidant response pathway.

Conformations of voltage-sensing domain III differentially define NaV channel closed- and open-state inactivation

Voltage-gated Na+ (NaV) channels underlie the initiation and propagation of action potentials (APs). Rapid inactivation after NaV channel opening, known as open-state inactivation, plays a critical role in limiting the AP duration. However, NaV channel inactivation can also occur before opening, namely closed-state inactivation, to tune the cellular excitability. The voltage-sensing domain (VSD) within repeat IV (VSD-IV) of the pseudotetrameric NaV channel α-subunit is known to be a critical regulator of NaV channel inactivation. Yet, the two processes of open- and closed-state inactivation predominate at different voltage ranges and feature distinct kinetics. How inactivation occurs over these different ranges to give rise to the complexity of NaV channel dynamics is unclear. Past functional studies and recent cryo-electron microscopy structures, however, reveal significant inactivation regulation from other NaV channel components. In this Hypothesis paper, we propose that the VSD of NaV repeat III (VSD-III), together with VSD-IV, orchestrates the inactivation-state occupancy of NaV channels by modulating the affinity of the intracellular binding site of the IFMT motif on the III-IV linker. We review and outline substantial evidence that VSD-III activates in two distinct steps, with the intermediate and fully activated conformation regulating closed- and open-state inactivation state occupancy by altering the formation and affinity of the IFMT crevice. A role of VSD-III in determining inactivation-state occupancy and recovery from inactivation suggests a regulatory mechanism for the state-dependent block by small-molecule anti-arrhythmic and anesthetic therapies.

Does the wall sound different? Variable acoustics in rehearsal rooms using small resonator structures in an acoustic panel.

As the name already states, multipurpose rooms are often used from various people for different intentions like meetings or musical practicing. One example are musical rehearsal rooms, where the acoustic specifications have to meet the requirements of musicians playing different instrument groups. To meet the desire for variable acoustics in a rehearsal room, musicians often like to adjust the room to there personal preferences, what is mostly done by adjusting the frequency dependent room decay curve (T60). Hence, a variable acoustic panel has been developed which consists of several small adjustable resonator structures. In a closed state, the structure acts like a resonator. Although Helmholtz resonators are mostly used at low frequencies, the acoustic panel can address acoustic resonance absorption in the mid-frequency range between 500 Hz and 1.500 Hz. The paper highlights especially the dimensioning of the resonators and its measurements in an impedance tube, a reverberation cabin and a reverberation room. Finally, the prototype of the acoustic panel has been analysed in different rehearsal rooms where musicians examine the panel and T60 differences between the open and closed state of the panel were measured.

OPEN and CLOSED state of SPIKE SARS-COV-2: relationship with some integrin binding. A biological molecular approach to better understand the coagulant effect

Related the physio-pathological process of COVID-19 disease it is interesting to focus to the aspect. Played by interaction of Sars-Cov-2 protein with integrins of human epithelial pulmonary cell. A bio molecular approach help in to deeply verify the involved factors and the results of this Activation RGD mediated. Of Great interest also the relationship with some vaccine strategy followed by the various pharmaceutical industry. The results of this work will be useful to think modification in some vaccine increasing the global safety and related some rare ADR.

Dynamic Shape Transformation of a DNA Scaffold Applied for an Enzyme Nanocarrier

Structural programmability and accurate addressability of DNA nanostructures are ideal characteristics for the platform of arranging enzymes with the nanoscale precision. In this study, a three-dimensional DNA scaffold was designed to enable a dynamic shape transition from an open plate-like structure to its closed state of a hexagonal prism structure. The two domains in the open state were folded together to transform into the closed state by hybridization of complementary short DNA closing keys at both of the facing edges in over 90% yield. The shape transformation of the DNA scaffold was extensively studied by means of the fluorescence energy transfer measurement, atomic force microscope images, and agarose gel electrophoretic analyses. A dimeric enzyme xylitol dehydrogenase was assembled on the DNA scaffold in its open state in a high-loading yield. The enzyme loaded on the scaffold was subsequently transformed to its closed state by the addition of short DNA closing keys. The enzyme encapsulated in the closed state displayed comparable activity to that in the open state, ensuring that the catalytic activity of the enzyme was well maintained in the DNA nanocarrier. The nanocarrier with efficient encapsulation ability is potentially applicable for drug delivery, biosensing, biocatalytic, and diagnostic tools.