Biochemical and mutational analysis of coenzyme B12 biosynthesis

The B12 coenzymes are required cofactors for at least 15 different enzymes that are widely distributed in nature and are essential for human health. Characterizing enzymes in each step of vitamin B12 metabolism in humans will provide important information for human B12-related disease diagnosis and therapy. Human ATP:cob(I)alamin adenosyltransferase (hATR) catalyzes the final step in the conversion of vitamin B12 into the active coenzyme, adenosylcobalamin. Inherited defects in this gene result in a rare but life-threatening disease, methylmalonyl aciduria. We conducted a random mutagenesis of the hATR coding sequence. Fifty-seven missense mutations were isolated mapping to 30 positions, 25 of which had not previously been shown to impair enzyme activity. Kinetic analysis and in vivo tests for enzyme activity were performed on the hATR variants, and mutations were mapped onto a hATR structural model. These studies functionally defined the hATR active site and tentatively implicated three amino acid residues in facilitating the reduction of cob(II)alamin to cob(I)alamin which is a prerequisite to adenosylation. Human cannot synthesize coenzyme B12 de novo but assimilate B12 complex precursors, which are only from certain prokaryotes. The biosynthesis of B12 has been studied extensively in Salmonella enterica, however, the function of PduX gene involved in B12-dependent propanediol degradation was still unknown. In vitro test including 31P NMR spectroscopy and enzyme assays established that purified PduX-His6 catalyzed the conversion of L-threonine and ATP to L-threonine-O-3-phosphate and ADP. A series of in vivo growth studies showed that PduX is an L-threonine kinase used for AdoCbl synthesis. The PduX enzyme belongs to GHMP kinase family by sequence similarity. Kinetic analysis indicated an ordered ternary complex mechanism in which ATP is the first substrate to bind. The lack of obvious solvent pH and isotope effects indicated that proton transfer is unlikely to be involved directly in the rate determining step of catalysis. Based on a multiple sequence alignment of PduX homologues and other GHMP family members, 14 PduX variant were constructed by site-directed mutagenesis. Comparison of the Circular Dichroism spectra and kinetic properties of the PduX variants to those of the wild-type enzyme functionally defined the L-threonine kinase active site

The Application of Cell-Free Protein Synthesis in Genetic Code Expansion for Post-translational Modifications

The translation system is a sophisticated machinery that synthesizes proteins from 20 canonical amino acids. Recently, the repertoire of such composition has been expanded by the introduction of non-canonical amino acids (ncAAs) with the genetic code expansion strategy, which provides proteins with designed properties and structures for protein studies and engineering. Although the genetic code expansion strategy has been mostly implemented by using living cells as the host, a number of limits such as poor cellular uptake or solubility of specific ncAA substrates and the toxicity of target proteins have hindered the production of certain ncAA-modified proteins. To overcome those challenges, cell-free protein synthesis (CFPS) has been applied as it allows the precise control of reaction components. Several approaches have been recently developed to increase the purity and efficiency of ncAA incorporation in CFPS. Here, we summarized recent development of CFPS with an emphasis on its applications in generating site-specific protein post-translational modifications by the genetic code expansion strategy

Characteristics and metabolic potential of biliary microbiota in patients with giant common bile duct stones

BackgroundEndoscopic retrograde cholangiopancreatography (ERCP) is an effective minimally invasive operation for the management of choledocholithiasis, while successful extraction is hampered by large diameter of stones. Emerging studies have revealed the close correlation between biliary microbiota and common bile duct stones (CBDS). In this study, we aimed to investigate the community characteristics and metabolic functions of biliary microbiota in patients with giant CBDS.MethodsEligible patients were prospectively enrolled in this study in First Affiliated Hospital of Soochow University from February 2022 to October 2022. Bile samples were collected through ERCP. The microbiota was analyzed using 16S rRNA sequencing. Metabolic functions were predicted by PICRUSTs 2.0 calculation based on MetaCyc database. Bile acids were tested and identified using ultra performance liquid chromatography-tandem mass spectrometry.ResultsA total of 26 patients were successfully included into final analysis, 8 in giant stone (GS) group and 18 in control group. Distinct biliary microbial composition was identified in patients with giant CBDS, with a significantly higher abundance of Firmicutes at phylum level. The unique composition at genus level mainly consisted of Enterococcus, Citrobacter, Lactobacillus, Pyramidobacter, Bifidobacterium and Shewanella. Pyramidobacter was exclusively found in GS group, along with the absence of Robinsoniella and Coprococcus. The contents of free bile acids were significantly higher in GS group, including cholic acid (98.39μmol/mL vs. 26.15μmol/mL, p=0.035), chenodesoxycholic acid (54.69μmol/mL vs. 5.86μmol/mL, p=0.022) and ursodeoxycholic acid (2.70μmol/mL vs. 0.17μmol/mL, p=0.047). Decreasing tendency of conjugated bile acids were also observed. Metabolic pathways concerning cholelithiasis were abundant in GS group, including geranylgeranyl diphosphate biosynthesis, gluconeogenesis, glycolysis and L-methionine biosynthesis.ConclusionsThis study demonstrated the community structure and metabolic potential of biliary microbiota in patients with giant CBDS. The unique biliary microbial composition holds valuable predictive potential for clinical conditions. These findings provide new insights into the etiology of giant CBDS from the perspective of biliary microbiota

From teleoperation to autonomous robot-assisted microsurgery: A survey

Robot-assisted microsurgery (RAMS) has many benefits compared to traditional microsurgery. Microsurgical platforms with advanced control strategies, high-quality micro-imaging modalities and micro-sensing systems are worth developing to further enhance the clinical outcomes of RAMS. Within only a few decades, microsurgical robotics has evolved into a rapidly developing research field with increasing attention all over the world. Despite the appreciated benefits, significant challenges remain to be solved. In this review paper, the emerging concepts and achievements of RAMS will be presented. We introduce the development tendency of RAMS from teleoperation to autonomous systems. We highlight the upcoming new research opportunities that require joint efforts from both clinicians and engineers to pursue further outcomes for RAMS in years to come

Preparation and enhanced electrorheological properties of elastomers filled with rod-shaped TiO2 particles

The morphology of dispersed particles has been proven to have a significant impact on performance of electrorheological (ER) materials, while there is a lack of relevant research on its impact on the properties of electrorheological elastomers (EREs). In this study, the TiO2 particles with spherical, short rod, and long rod shape were fabricated with sol-gel method, and the EREs were prepared with these three kinds of particles as dispersion phase. Particle characterization results show that the rod-shape TiO2 particles with larger average size exhibit a combination of anatase and brookite phase. The viscoelastic properties of three types of EREs under varying strain amplitude and shear frequency were tested. The results indicate that the long rod-shape TiO2 particles filled EREs shows higher storage modulus G′ and higher relative ER effect within the electric field from 0 to 3 kV/mm. The observations indicate the use of rod-shape TiO2 particles in the form of brookite phase may help enhance the ER properties of elastomers. The investigation contributes to the designing, preparation, and application of anisotropic ERE

One-shot domain-adaptive imitation learning via progressive learning applied to robotic pouring

Traditional deep learning-based visual imitation learning techniques require a large amount of demonstration data for model training, and the pre-trained models are difficult to adapt to new scenarios. To address these limitations, we propose a unified framework using a novel progressive learning approach comprised of three phases: i) a coarse learning phase for concept representation, ii) a fine learning phase for action generation, and iii) an imaginary learning phase for domain adaptation. Overall, this approach leads to a one-shot domain-adaptive imitation learning framework. We use robotic pouring as an example task to evaluate its effectiveness. Our results show that the method has several advantages over contemporary end-to-end imitation learning approaches, including an improved success rate for task execution and more efficient training for deep imitation learning. In addition, the generalizability to new domains is improved, as demonstrated here with novel backgrounds, target containers, and granule combinations in the experiment. We believe that the proposed method is broadly applicable to various industrial or domestic applications that involve deep imitation learning for robotic manipulation, and where the target scenarios are diverse and human demonstration data is limited. For project video, please check our website:. Note to Practitioners —The motivation of this paper is to develop a progressive learning framework, which can be used for both service and industrial robots to learn from human demonstrations, and then transfer the learned skill to different scenarios with ease. We use the robotic pouring task as an example to demonstrate the effectiveness of our proposed method, since pouring is an essential skill for service robots to assist humans’ daily lives, and can benefit robot automation in wet-lab industries. The aim of this research is to enable robots to obtain visuomotor skills (such as the pouring skill), and accomplish the tasks with a high success rate using our proposed progressive learning method. We conducted experiments to show that the proposed method has good performance, high data efficiency and evident generalizability. This is significant for intelligent robots working in various practical applications

Mg3(Bi,Sb)2 single crystals towards high thermoelectric performance

The rapid growth of the thermoelectric cooler market makes the development of novel room temperature thermoelectric materials of great importance. Ternary n-type Mg3(Bi,Sb)2 alloys are promising alternatives to the state-of-the-art Bi2(Te,Se)3 alloys but grain boundary resistance is the most important limitation. n-type Mg3(Bi,Sb)2 single crystals with negligible grain boundaries are expected to have particularly high zT but have rarely been realized due to the demanding Mg-rich growth conditions required. Here, we report, for the first time, the thermoelectric properties of n-type Mg3(Bi,Sb)2 alloyed single crystals grown by a one-step Mg-flux method using sealed tantalum tubes. High weighted mobility ∼140 cm2 V−1 s−1 and a high zT of 0.82 at 315 K are achieved in Y-doped Mg3Bi1.25Sb0.75 single crystals. Through both experimental angle-resolved photoemission spectroscopy and theoretical calculations, we denote the origin of the high thermoelectric performance from a point of view of band widening effect and electronegativity, as well as the necessity to form high Bi/Sb ratio ternary Mg3(Bi,Sb)2 alloys. The present work paves the way for further development of Mg3(Bi,Sb)2 for near room temperature thermoelectric applications

Thermoelectric Properties of Novel Semimetals: A Case Study of YbMnSb2

The emerging class of topological materials provides a platform to engineer exotic electronic structures for a variety of applications. As complex band structures and Fermi surfaces can directly benefit thermoelectric performance it is important to identify the role of featured topological bands in thermoelectrics particularly when there are coexisting classic regular bands. In this work, the contribution of Dirac bands to thermoelectric performance and their ability to concurrently achieve large thermopower and low resistivity in novel semimetals is investigated. By examining the YbMnSb2 nodal line semimetal as an example, the Dirac bands appear to provide a low resistivity along the direction in which they are highly dispersive. Moreover, because of the regular-band-provided density of states, a large Seebeck coefficient over 160 µV K−1 at 300 K is achieved in both directions, which is very high for a semimetal with high carrier concentration. The combined highly dispersive Dirac and regular bands lead to ten times increase in power factor, reaching a value of 2.1 mW m−1 K−2 at 300 K. The present work highlights the potential of such novel semimetals for unusual electronic transport properties and guides strategies towards high thermoelectric performance. © 2020 The Authors. Advanced Materials published by Wiley-VCH Gmb

Destructive quantum interference in <i>meta</i>-oligo(phenyleneethynylene) molecular wires with gold-graphene heterojunctions.

Quantum interference (QI) is well recognised as a significant contributing factor to the magnitude of molecular conductance values in both single-molecule and large area junctions. Numerous structure-property relationship studies have shown that para-connected oligo(phenyleneethynylene) (OPE) based molecular wires exemplify the impact of constructive quantum interference (CQI), whilst destructive quantum interference (DQI) effects are responsible for the orders of magnitude lower conductance of analogous meta-contacted OPE derivatives, despite the somewhat shorter effective tunnelling distance. Since molecular conductance is related to the value of the transmission function, evaluated at the electrode Fermi energy, T(EF), which in turn is influenced by the presence and relative energy of (anti)resonances, it follows that the relative single-molecule conductance of para- and meta-contacted OPE-type molecules is tuned both by the anchor group and the nature of the electrode materials used in the construction of molecular junctions (gold|molecule|gold vs. gold|molecule|graphene). It is shown here that whilst amine-contacted junctions show little influence of the electrode material on molecular conductance due to the similar electrode-molecule coupling through this anchor group to both types of electrodes, the weaker coupling between thiomethyl and ethynyl anchors and the graphene substrate electrode results in a relative enhancement of the DQI effect. This work highlights an additional parameter space to explore QI effects and establishes a new working model based on the electrode materials and anchor groups in modulating QI effects beyond the chemical structure of the molecular backbone