Key residues in the nicotinic acetylcholine receptor β2 subunit contribute to α-conotoxin LvIA binding

alpha-Conotoxin LvIA (alpha-CTx LvIA) is a small peptide from the venom of the carnivorous marine gastropod Conus lividus and is the most selective inhibitor of alpha 3 beta 2 nicotinic acetylcholine receptors (nAChRs) known to date. It can distinguish the alpha 3 beta 2 nAChR subtype from the alpha 6 beta 2* (*indicates the other subunit) and alpha 3 beta 4 nAChR subtypes. In this study, we performed mutational studies to assess the influence of residues of the beta 2 subunit versus those of the beta 4 subunit on the binding of alpha-CTx LvIA. Although two beta 2 mutations, alpha 3 beta 2[F119Q] and alpha 3 beta 2[T59K], strongly enhanced the affinity of LvIA, the beta 2 mutation alpha 3 beta 2[V111I] substantially reduced the binding of LvIA. Increased activity of LvIA was also observed when the beta 2-T59L mutant was combined with the alpha 3 subunit. There were no significant difference in inhibition of alpha 3 beta 2[T59I], alpha 3 beta 2[Q34A], and alpha 3 beta 2[K79A] nAChRs when compared with wild-type alpha 3 beta 2 nAChR. alpha-CTx LvIA displayed slower off-rate kinetics at alpha 3 beta 2[F119Q] and alpha 3 beta 2[T59K] than at the wild-type receptor, with the latter mutant having the most pronounced effect. Taken together, these data provide evidence that the beta 2 subunit contributes to alpha-CTx LvIA binding and selectivity. The results demonstrate that Val(111) is critical and facilitates LvIA binding; this position has not previously been identified as important to binding of other 4/7 framework alpha-conotoxins. Thr(59) and Phe(119) of the beta 2 subunit appear to interfere with LvIA binding, and their replacement by the corresponding residues of the beta 4 subunit leads to increased affinity

ALMA High-resolution Spectral Survey of Thioformaldehyde (H2CS) Towards Massive Protoclusters

Investigating the temperature and density structures of gas in massive protoclusters is crucial for understanding the chemical properties therein. In this study, we present observations of the continuum and thioformaldehyde (H2CS) lines at 345 GHz of 11 massive protoclusters using the Atacama Large Millimeter/submillimeter Array (ALMA) telescope. High spatial resolution and sensitivity observations have detected 145 continuum cores from the 11 sources. H2CS line transitions are observed in 72 out of 145 cores, including line-rich cores, warm cores and cold cores. The H2 column densities of the 72 cores are estimated from the continuum emission which are larger than the density threshold value for star formation, suggesting that H2CS can be widely distributed in star-forming cores with different physical environments. Rotation temperature and column density of H2CS are derived by use of the XCLASS software. The results show the H2CS abundances increase as temperature rises and higher gas temperatures are usually associated with higher H2CS column densities. The abundances of H2CS are positively correlated with its column density, suggesting that the H2CS abundances are enhanced from cold cores, warm cores to line-rich cores in star forming regions.Comment: 23 pages, 7 figures, 6 tables, accepted by Ap

Characterization of a novel alpha-conotoxin TxID from Conus textile that potently blocks rat alpha3/beta4 nicotinic acetylcholine receptors

The alpha 3 beta 4 nAChRs are implicated in pain sensation in the PNS and addiction to nicotine in the CNS. We identified an alpha-4/6-conotoxin (CTx) TxID from Conus textile. The new toxin consists of 15 amino acid residues with two disulfide bonds. TxID was synthesized using solid phase methods, and the synthetic peptide was functionally tested on nAChRs heterologously expressed in Xenopus laevis oocytes. TxID blocked rat alpha 3 beta 4 nAChRs with a 12.5 nM IC50, which places it among the most potent alpha 3 beta 4 nAChR antagonists. TxID also blocked the closely related alpha 6/alpha 3 beta 4 with a 94 nM IC50 but showed little activity on other nAChR subtypes. NMR analysis showed that two major structural isomers exist in solution, one of which adopts a regular alpha-CTx fold but with different surface charge distribution to other 4/6 family members. alpha-CTx TxID is a novel tool with which to probe the structure and function of alpha 3 beta 4 nAChRs

Effects of pore structure characteristics on performance of sintered bi-porous Ti3AlC2 wicks

Loop heat pipe, carried out by liquid-gas phase transition, has been attracted as an efficient heat management device in high-heat-flux, long-distance and anti-gravity situations. The capillary pressure provided by the porous wick in the evaporator drives circulation for working liquid, affecting the heat exchange efficiency for loop heat pipes directly. In this work, the bi-porous Ti _3 AlC _2 wick was produced by reaction sintering and pore formers dissolution and porosity and pore size were regulated by varying pore formers content and cold pressure. The pore size distribution was characterized by mercury intrusion and effects on capillary performance and thermal conductivity were also analyzed. The porosity and proportion of large pores increased with the increased pore formers content, causing the enhancement of capillary performance and the reduction of thermal conductivity. As the cold pressure increased, pore size distribution concentrated, the proportion of fine pores increased and the porosity fluctuated slightly, increasing the capillary pressure and improving capillary performance and thermal conductivity. Based on experimental results, the parameter of pore size proportion was proposed to modify the Alexander model to predict the thermal conductivity of porous materials accurately, guide the design for pore structure and promote the transfer capacity

Laser Welding of TA15 Titanium Alloy and Inconel 718 Dissimilar Metals

This paper studied the effect of laser welding technology on dissimilar metal welding joints of TA15 titanium alloy and Inconel 718 nickel-based alloy. The research results indicate that the laser welding of TA15 titanium alloy and Inconel 718 nickel-based alloy directly was difficult to form well, which due to the intermetallic compounds caused the joint brittle. When the pure Cu foil was used as the filling layer, the quality of the welding joints can be improved effectively. The experimental results also indicate that there were brittle intermetallic-compounds in the laser welding seam, and the laser power had an important influence on the performance and mechanical properties of the dissimilar metal joint. The maximum average tensile strength of the welding joint of 2300 W was increased to 252.32 MPa. Scanning electron microscope(SEM) results show that the fracture morphology was river pattern, a typical morphological of cleavage fracture, and the mode was brittle fracture

A Novel α4/7-Conotoxin QuIA Selectively Inhibits α3β2 and α6/α3β4 Nicotinic Acetylcholine Receptor Subtypes with High Efficacy

α6β4 nAChR is expressed in the peripheral and central nervous systems and is associated with pain, addiction, and movement disorders. Natural α-conotoxins (α-CTxs) can effectively block different nAChR subtypes with higher efficacy and selectivity. However, the research on α6β4 nAChR is relatively poor, partly because of the lack of available target-specific α-CTxs. In this study, we synthesized a novel α-4/7 conotoxin QuIA that was found from Conus quercinus. We investigated the efficacy of this peptide to different nAChR subtypes using a two-electrode voltage-clamp technique. Remarkably, we found α-QuIA inhibited the neuronal α3β2 and α6/α3β4 nAChR subtypes with significantly high affinity (IC50 was 55.7 nM and 90.68 nM, respectively), and did not block other nAChR subtypes even at a high concentration of 10 μM. In contrast, most α-CTxs have been determined so far to effectively block the α6/α3β4 nAChR subtype while also maintaining a similar higher efficacy against the closely related α6β2β3 and/or α3β4 subtypes, which are different from QuIA. In conclusion, α-QuIA is a novel α4/7-CTx, which has the potential to develop as an effective neuropharmacology tool to detect the function of α6β4 nAChR

Engineering Enhanced Antimicrobial Properties in α-Conotoxin RgIA through D-Type Amino Acid Substitution and Incorporation of Lysine and Leucine Residues

Antimicrobial peptides (AMPs), acknowledged as host defense peptides, constitute a category of predominant cationic peptides prevalent in diverse life forms. This study explored the antibacterial activity of α-conotoxin RgIA, and to enhance its stability and efficacy, D-amino acid substitution was employed, resulting in the synthesis of nine RgIA mutant analogs. Results revealed that several modified RgIA mutants displayed inhibitory efficacy against various pathogenic bacteria and fungi, including Candida tropicalis and Escherichia coli. Mechanistic investigations elucidated that these polypeptides achieved antibacterial effects through the disruption of bacterial cell membranes. The study further assessed the designed peptides’ hemolytic activity, cytotoxicity, and safety. Mutants with antibacterial activity exhibited lower hemolytic activity and cytotoxicity, with Pep 8 demonstrating favorable safety in mice. RgIA mutants incorporating D-amino acids exhibited notable stability and adaptability, sustaining antibacterial properties across diverse environmental conditions. This research underscores the potential of the peptide to advance innovative oral antibiotics, offering a novel approach to address bacterial infections

Optimal Cleavage and Oxidative Folding of α-Conotoxin TxIB as a Therapeutic Candidate Peptide

Alpha6beta2 nicotinic acetylcholine receptors (nAChRs) are potential therapeutic targets for the treatment of several neuropsychiatric diseases, including addiction and Parkinson’s disease. Alpha-conotoxin (α-CTx) TxIB is a uniquely selective ligand, which blocks α6/α3β2β3 nAChRs only, but does not block the other subtypes. Therefore, α-CTx TxIB is a valuable therapeutic candidate peptide. Synthesizing enough α-CTx TxIB with high yield production is required for conducting wide-range testing of its potential medicinal applications. The current study optimized the cleavage of synthesized α-CTx TxIB resin-bounded peptide and folding of the cleaved linear peptide. Key parameters influencing cleavage and oxidative folding of α-CTx TxIB were examined, such as buffer, redox agents, pH, salt, co-solvent and temperature. Twelve conditions were used for cleavage optimization. Fifty-four kinds of one-step oxidative solution were used to assess their effects on each α-CTx TxIB isomers’ yield. The result indicated that co-solvent choices were particularly important. Completely oxidative folding of globular isomer was achieved when the NH4HCO3 or Tris-HCl folding buffer at 4 °C contained 40% of co-solvent DMSO, and GSH:GSSG (2:1) or GSH only with pH 8~8.7

YOLO-Weld: A Modified YOLOv5-Based Weld Feature Detection Network for Extreme Weld Noise

Weld feature point detection is a key technology for welding trajectory planning and tracking. Existing two-stage detection methods and conventional convolutional neural network (CNN)-based approaches encounter performance bottlenecks under extreme welding noise conditions. To better obtain accurate weld feature point locations in high-noise environments, we propose a feature point detection network, YOLO-Weld, based on an improved You Only Look Once version 5 (YOLOv5). By introducing the reparameterized convolutional neural network (RepVGG) module, the network structure is optimized, enhancing detection speed. The utilization of a normalization-based attention module (NAM) in the network enhances the network’s perception of feature points. A lightweight decoupled head, RD-Head, is designed to improve classification and regression accuracy. Furthermore, a welding noise generation method is proposed, increasing the model’s robustness in extreme noise environments. Finally, the model is tested on a custom dataset of five weld types, demonstrating better performance than two-stage detection methods and conventional CNN approaches. The proposed model can accurately detect feature points in high-noise environments while meeting real-time welding requirements. In terms of the model’s performance, the average error of detecting feature points in images is 2.100 pixels, while the average error in the world coordinate system is 0.114 mm, sufficiently meeting the accuracy needs of various practical welding tasks

Discovery Methodology of Novel Conotoxins from <i>Conus</i> Species

Cone snail venoms provide an ideal resource for neuropharmacological tools and drug candidates discovery, which have become a research hotspot in neuroscience and new drug development. More than 1,000,000 natural peptides are produced by cone snails, but less than 0.1% of the estimated conotoxins has been characterized to date. Hence, the discovery of novel conotoxins from the huge conotoxin resources with high-throughput and sensitive methods becomes a crucial key for the conotoxin-based drug development. In this review, we introduce the discovery methodology of new conotoxins from various Conus species. It focuses on obtaining full N- to C-terminal sequences, regardless of disulfide bond connectivity through crude venom purification, conotoxin precusor gene cloning, venom duct transcriptomics, venom proteomics and multi-omic methods. The protocols, advantages, disadvantages, and developments of different approaches during the last decade are summarized and the promising prospects are discussed as well