RESUMEN
Methionyl-tRNA synthetase (MetRS) catalyzes the attachment of l-methionine (l-Met) to tRNAMet to generate methionyl-tRNAMet, an essential substrate for protein translation within ribosome. Owing to its indispensable biological function and the structural discrepancies with human counterpart, bacterial MetRS is considered an ideal target for developing antibacterials. Herein, chlorhexidine (CHX) was identified as a potent binder of Staphylococcus aureus MetRS (SaMetRS) through an ATP-aided affinity screening. The co-crystal structure showed that CHX simultaneously occupies the enlarged l-Met pocket (EMP) and the auxiliary pocket (AP) of SaMetRS with its two chlorophenyl groups, while its central hexyl linker swings upwards to interact with some conserved hydrophobic residues. ATP adopts alternative conformations in the active site cavity, and forms ionic bonds and water-mediated hydrogen bonds with CHX. Consistent with this synergistic binding mode, ATP concentration-dependently enhanced the binding affinity of CHX to SaMetRS from 10.2 µM (no ATP) to 0.45 µM (1 mM ATP). While it selectively inhibited two representative type 1 MetRSs from S. aureus and Enterococcus faecalis, CHX did not show significant interactions with three tested type 2 MetRSs, including human cytoplasmic MetRS, in the enzyme inhibition and biophysical binding assays, probably due to the conformational differences between two types of MetRSs at their EMP and AP. Our findings on CHX may inspire the design of MetRS-directed antimicrobials in future.
Asunto(s)
Metionina-ARNt Ligasa , Humanos , Metionina-ARNt Ligasa/química , Metionina-ARNt Ligasa/genética , Metionina-ARNt Ligasa/metabolismo , Clorhexidina/farmacología , Staphylococcus aureus , ARN de Transferencia de Metionina/metabolismo , Bacterias Grampositivas/metabolismo , Adenosina Trifosfato/metabolismoRESUMEN
The role of co-agonists of glucagon-like peptide-1 receptor (GLP-1R) and glucagon receptor (GCGR) in chronic kidney disease (CKD) remains unclear. Herein we found that GLP-1R and GCGR expression levels were lower in the kidneys of mice with CKD compared to healthy mice and were correlated with disease severity. Interestingly, GLP-1R or GCGR knockdown aggravated the progression of kidney injury in both diabetic db/db mice and non-diabetic mice undergoing unilateral ureteral obstruction (UUO). Based on the importance of GLP-1R and GCGR in CKD, we reported a novel monomeric peptide, 1907-B, with dual-agonism on both GLP-1R and GCGR. The data confirmed that 1907-B had a longer half-life than long-acting semaglutide in rats or cynomolgus monkeys (â¼2-3 fold) and exhibited better therapeutic contribution to CKD than best-in-class monoagonists, semaglutide, or glucagon, in db/db mice and UUO mice. Various lock-of-function models, including selective pharmacological activation and genetic knockdown, confirmed that 1907-B's effects on ameliorating diabetic nephropathy in db/db mice, as well as inhibiting kidney fibrosis in UUO mice, were mediated through GLP-1 and glucagon signaling. These findings highlight that 1907-B, a novel GLP-1R and GCGR co-agonist, exerts multifactorial improvement in kidney injuries and is an effective and promising therapeutic option for CKD treatment.
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With the advent of the sixth-generation mobile communication standard (6â G), the visible light communication (VLC) technology based on wavelength division multiplexing (WDM) technology can effectively solve the problem of shortage of spectrum resources and insufficient channel capacity. This paper introduces one of our technical achievements, namely the construction of a near-real-time visible light laser communication (VLLC) system based on WDM, which includes a self-designed 10-λ fully-packaged visible light laser emission module, 1 m multimode fiber - 0.175 m free space - 1 m multimode fiber optical transmission link, and receiver array. In the transmitter system, we adopt adaptive discrete multitone (DMT) modulation technique combined with Quadrature Amplitude Modulation (QAM) modulation scheme to obtain maximum spectral efficiency (SE). In the receiving system, we utilize the sparse-structured reservoir computing post-equalization algorithm to achieve superior equalization performance on the basis of the traditional post-equalization algorithm. The experimental results indicate that this quasi-real-time communication system has achieved a signal transmission rate of 113.175Gbps. To the best of our knowledge, this work has set a record in the field of high-speed visible light laser communication. Therefore, the laser communication system constructed by this work, with its flexibility in deployment and high-speed performance, demonstrates the significant potential application of visible light laser communication in data center interconnection and high-speed indoor access networks.
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In this paper, we studied a series of high-speed photodetectors (PD) with different super-lattice interlayer periods and the scale of the effective area to examine their communication performance. The mini-PDs are designed with a single 1 mm × 1 mm effective area. The mini-PDs have three different super-lattice (SL) periods in the interlayer: 8, 15, and 32. The micro-PD sample has multiple 50um by 50um photosensitive areas that form a 4 × 4 receiver array, which shares a common N electrode. Its SL period is 26. The experiment shows that mini-PDs have the advantages such as better tolerance to beam spot deviation, larger field of view (FoV), higher responsibility, and wider peak width in spectral response. But micro-LED samples outperform the others in communication capacity and wavelength selectivity. The 8, 15, and 32 SL mini-PD samples achieve 6.6, 7.3, and 8.8 Gb/s data rates, respectively. The micro-PD gains the maximum data rate of 14.38Gb/s without applying waveform level post-equalization, and 15.26Gb/s after using an NN-based post-equalizer. This experiment shows that with proper DSP, GaN-based PD would be suitable for high-speed VLC systems, especially for the short wavelength spectrum in visible light.
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This Letter presents an experimental demonstration of a visible light communication system utilizing a LiNbO3 external modulator to support the transmission of pulse amplitude modulation (PAM)-4 signals. To solve the problem of the low-frequency fluctuations and inter-symbol interference (ISI) introduced by the external modulator-based system, a neural network with a low-frequency signal as the second label (LFNN) is proposed. A data rate of 8.8 Gbps using PAM-4 is experimentally achieved under the 7% hard-decision forward error correction (HD-FEC) bit-error-ratio (BER) limit of 3.8 × 10-3. To the best of our knowledge, this work represents the highest transmission data rate achieved thus far using external modulation in visible light communication systems.
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This Letter proposes a novel, to the best of our knowledge, approach utilizing a delta-sigma modulation (DSM)-based 1-bit autoencoder (AE) for efficient encoding and decoding in various channel conditions. Simulation analysis demonstrates the AE's ability to mitigate noise by reducing a peak-to-average power ratio (PAPR) and enhancing an in-band power of the signals, particularly under low signal-to-noise ratios (SNRs). The AE-DSM achieves theoretical transmission performance even at SNRs below 6â dB. In a 40-m free-space link experiment, the AE-DSM exhibits an 8.4-dB lower bit error rate (BER) compared to 64QAM-DSM, enabling a transmission rate of 1.31â Gbps. Furthermore, the 1-bit AE-DSM significantly reduces power consumption in the receiving analog-to-digital converter (ADC), facilitates transmission at low SNRs, and effectively mitigates nonlinear effects. Consequently, the DSM-based AE holds immense potential for future mobile fronthaul links.
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Inverse design has been widely studied as an efficient method to reduce footprint and improve performance for integrated silicon photonic (SiP) devices. In this study, we have used inverse design to develop a series of ultra-compact dual-band wavelength demultiplexing power splitters (WDPSs) that can simultaneously perform both wavelength demultiplexing and 1:1 optical power splitting. These WDPSs could facilitate the potential coexistence of dual-band passive optical networks (PONs). The design is performed on a standard silicon-on-insulator (SOI) platform using, what we believe to be, a novel two-step direct binary search (TS-DBS) method and the impact of different hyperparameters related to the physical structure and the optimization algorithm is analyzed in detail. Our inverse-designed WDPS with a minimum feature size of 130â nm achieves a 12.77-times reduction in footprint and a slight increase in performance compared with the forward-designed WDPS. We utilize the optimal combination of hyperparameters to design another WDPS with a minimum feature size reduced to 65â nm, which achieves ultra-low insertion losses of 0.36â dB and 0.37â dB and crosstalk values of -19.91â dB and -17.02â dB at wavelength channels of 1310â nm and 1550â nm, respectively. To the best of our knowledge, the hyperparameters of optimization-based inverse design are systematically discussed for the first time. Our work demonstrates that appropriate setting of hyperparameters greatly improves device performance, throwing light on the manipulation of hyperparameters for future inverse design.
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Tryptophanyl-tRNA synthetase (TrpRS) links tryptophan to tRNATrp, thereby playing an indispensable role in protein translation. Unlike most class I aminoacyl-tRNA synthetases (AARSs), TrpRS functions as a homodimer. Herein, we captured an 'open-closed' asymmetric structure of Escherichia coli TrpRS (EcTrpRS) with one active site occupied by a copurified intermediate product and the other remaining empty, providing structural evidence for the long-discussed half-of-the-sites reactivity of bacterial TrpRS. In contrast to its human counterpart, bacterial TrpRS may rely on this asymmetric conformation to functionally bind with substrate tRNA. As this asymmetric conformation is probably a dominant form of TrpRS purified from bacterial cells, we performed fragment screening against asymmetric EcTrpRS to support antibacterial discovery. Nineteen fragment hits were identified, and 8 of them were successfully cocrystallized with EcTrpRS. While a fragment named niraparib bound to the L-Trp binding site of the 'open' subunit, the other 7 fragments all bound to an unprecedented pocket at the interface between two TrpRS subunits. Binding of these fragments relies on residues specific to bacterial TrpRS, avoiding undesired interactions with human TrpRS. These findings improve our understanding of the catalytic mechanism of this important enzyme and will also facilitate the discovery of bacterial TrpRS inhibitors with therapeutic potential.
Asunto(s)
Antiinfecciosos , Proteínas de Escherichia coli , Escherichia coli , Triptófano-ARNt Ligasa , Sitios de Unión , Dominio Catalítico , Triptófano/metabolismo , Triptófano-ARNt Ligasa/genética , Escherichia coli/enzimología , Escherichia coli/genética , Proteínas de Escherichia coli/genéticaRESUMEN
As a class of essential enzymes in protein translation, aminoacyl-transfer RNA (tRNA) synthetases (aaRSs) are organized into two classes of 10 enzymes each, based on two conserved active site architectures. The (αß)2 glycyl-tRNA synthetase (GlyRS) in many bacteria is an orphan aaRS whose sequence and unprecedented X-shaped structure are distinct from those of all other aaRSs, including many other bacterial and all eukaryotic GlyRSs. Here, we report a cocrystal structure to elucidate how the orphan GlyRS kingdom specifically recognizes its substrate tRNA. This structure is sharply different from those of other aaRS-tRNA complexes but conforms to the clash-free, cross-class aaRS-tRNA docking found with conventional structures and reinforces the class-reconstruction paradigm. In addition, noteworthy, the X shape of orphan GlyRS is condensed with the largest known spatial rearrangement needed by aaRSs to capture tRNAs, which suggests potential nonactive site targets for aaRS-directed antibiotics, instead of less differentiated hard-to-drug active site locations.
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Aminoacil-ARNt Sintetasas , Glicina-ARNt Ligasa , Glicina-ARNt Ligasa/genética , Glicina-ARNt Ligasa/química , Glicina-ARNt Ligasa/metabolismo , Aminoacil-ARNt Sintetasas/química , Aminoacil-ARNt Sintetasas/genética , Aminoacil-ARNt Sintetasas/metabolismo , Ligasas/metabolismo , ARN de Transferencia , Dominio CatalíticoRESUMEN
Aminoacyl-tRNA synthetases (aaRSs) are promising antimicrobial targets due to their essential roles in protein translation, and expanding their inhibitory mechanisms will provide new opportunities for drug discovery. We report here that halofuginone (HF), an herb-derived medicine, moderately inhibits prolyl-tRNA synthetases (ProRSs) from various pathogenic bacteria. A cocrystal structure of Staphylococcus aureus ProRS (SaProRS) with HF and an ATP analog was determined, which guided the design of new HF analogs. Compound 3 potently inhibited SaProRS at IC50 = 0.18 µM and Kd = 30.3 nM and showed antibacterial activities with an MIC of 1-4 µg/mL in vitro. The bacterial drug resistance to 3 only developed at a rate similar to or slower than those of clinically used antibiotics in vitro. Our study indicates that the scaffold and ATP-aided inhibitory mechanism of HF could apply to bacterial ProRS and also provides a chemical validation for using bacterial ProRS as an antibacterial target.
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Aminoacil-ARNt Sintetasas , Bacterias , ARN de Transferencia , Adenosina TrifosfatoRESUMEN
Currently, there is still no effective curative treatment for the development of late-stage liver fibrosis. Here, we have illustrated that TB001, a dual glucagon-like peptide-1 receptor/glucagon receptor (GLP-1R/GCGR) agonist with higher affinity towards GCGR, could retard the progression of liver fibrosis in various rodent models, with remarkable potency, selectivity, extended half-life and low toxicity. Four types of liver fibrosis animal models which were induced by CCl4, α-naphthyl-isothiocyanate (ANIT), bile duct ligation (BDL) and Schistosoma japonicum were used in our study. We found that TB001 treatment dose-dependently significantly attenuated liver injury and collagen accumulation in these animal models. In addition to decreased levels of extracellular matrix (ECM) accumulation during hepatic injury, activation of hepatic stellate cells was also inhibited via suppression of TGF-ß expression as well as downstream Smad signaling pathways particularly in CCl4-and S. japonicum-induced liver fibrosis. Moreover, TB001 attenuated liver fibrosis through blocking downstream activation of pro-inflammatory nuclear factor kappa B/NF-kappa-B inhibitor alpha (NFκB/IKBα) pathways as well as c-Jun N-terminal kinase (JNK)-dependent induction of hepatocyte apoptosis. Furthermore, GLP-1R and/or GCGR knock-down results represented GCGR played an important role in ameliorating CCl4-induced hepatic fibrosis. Therefore, TB001 can be used as a promising therapeutic candidate for the treatment of multiple causes of hepatic fibrosis demonstrated by our extensive pre-clinical evaluation of TB001.
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Aminoacyl-tRNA synthetases (aaRSs) are promising drug targets due to their essential roles in protein translation. Although current inhibitors primarily occupy one or two of the three substrate binding sites on aaRSs, we report here the structure-based design of the first class of triple-site aaRS inhibitors by targeting Salmonella enterica threonyl-tRNA synthetase (SeThrRS). Competition of our compounds with all three substrates on SeThrRS binding was confirmed via isothermal titration calorimetry assays. Cocrystal structures of three compounds bound to SeThrRS unambiguously confirmed their substrate-mimicking triple-site binding mode. Compound 36j exhibited the best enzyme activity against SeThrRS with IC50 = 19 nM and Kd = 35.4 nM. Compounds 36b, 36k, and 36l exhibited antibacterial activities with minimum inhibitory concentration values of 2-8 µg/mL against the tested bacteria, which are superior to those of the reported dual-site ThrRS inhibitors. Our study provides the first proof-of-concept for developing triple-site inhibitors against aaRSs, inspiring future aaRS-based drug discoveries.
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Aminoacil-ARNt Sintetasas , Aminoacil-ARNt Sintetasas/metabolismo , Antibacterianos/química , Antibacterianos/farmacología , Descubrimiento de Drogas , Pruebas de Sensibilidad Microbiana , ARN de TransferenciaRESUMEN
Methionyl-tRNA synthetase (MetRS) charges tRNAMet with l-methionine (L-Met) to decode the ATG codon for protein translation, making it indispensable for all cellular lives. Many gram-positive bacteria use a type 1 MetRS (MetRS1), which is considered a promising antimicrobial drug target due to its low sequence identity with human cytosolic MetRS (HcMetRS, which belongs to MetRS2). Here, we report crystal structures of a representative MetRS1 from Staphylococcus aureus (SaMetRS) in its apo and substrate-binding forms. The connecting peptide (CP) domain of SaMetRS differs from HcMetRS in structural organization and dynamic movement. We screened 1049 chemical fragments against SaMetRS preincubated with or without substrate ATP, and ten hits were identified. Four cocrystal structures revealed that the fragments bound to either the L-Met binding site or an auxiliary pocket near the tRNA CCA end binding site of SaMetRS. Interestingly, fragment binding was enhanced by ATP in most cases, suggesting a potential ATP-assisted ligand binding mechanism in MetRS1. Moreover, co-binding with ATP was also observed in our cocrystal structure of SaMetRS with a class of newly reported inhibitors that simultaneously occupied the auxiliary pocket, tRNA site and L-Met site. Our findings will inspire the development of new MetRS1 inhibitors for fighting microbial infections.
Asunto(s)
Metionina-ARNt Ligasa , Humanos , Metionina-ARNt Ligasa/química , Ligandos , Sitios de Unión , Staphylococcus aureus/genética , Staphylococcus aureus/metabolismo , Metionina/metabolismo , Adenosina Trifosfato/metabolismoRESUMEN
AaRSs (aminoacyl-tRNA synthetases) group into two ten-member classes throughout evolution, with unique active site architectures defining each class. Most are monomers or homodimers but, for no apparent reason, many bacterial GlyRSs are heterotetramers consisting of two catalytic α-subunits and two tRNA-binding ß-subunits. The heterotetrameric GlyRS from Escherichia coli (EcGlyRS) was historically tested whether its α- and ß-polypeptides, which are encoded by a single mRNA with a gap of three in-frame codons, are replaceable by a single chain. Here, an unprecedented X-shaped structure of EcGlyRS shows wide separation of the abutting chain termini seen in the coding sequences, suggesting strong pressure to avoid a single polypeptide format. The structure of the five-domain ß-subunit is unique across all aaRSs in current databases, and structural analyses suggest these domains play different functions on α-subunit binding, ATP coordination and tRNA recognition. Moreover, the X-shaped architecture of EcGlyRS largely fits with a model for how two classes of tRNA synthetases arose, according to whether enzymes from opposite classes can simultaneously co-dock onto separate faces of the same tRNA acceptor stem. While heterotetrameric GlyRS remains the last structurally uncharacterized member of aaRSs, our study contributes to a better understanding of this ancient and essential enzyme family.
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Dominio Catalítico/genética , Escherichia coli/genética , Glicina-ARNt Ligasa/genética , ARN de Transferencia de Glicerina/química , Adenosina Trifosfato/metabolismo , Cristalografía por Rayos X , Glicina/química , Modelos Moleculares , ARN de Transferencia de Glicerina/genéticaRESUMEN
Staphylopine (StP) and other nicotianamine-like metallophores are crucial for many pathogens to acquire the transition metals from hosts during invasion. CntL from Staphylococcus aureus (SaCntL) catalyzes the condensation of the 2-aminobutyrate (Ab) moiety of S-adenosylmethionine (SAM) with D-histidine in the biosynthesis of StP. Here, we report the crystal structures of SaCntL in complex with either SAM or two products. The structure of SaCntL consists of an N-terminal four-helix bundle (holding catalytic residue E84) and a C-terminal Rossmann fold (binding the substrates). The sequence connecting the N- and C-terminal domains (N-C linker) in SaCntL was found to undergo conformational alternation between open and closed states. Our structural and biochemical analyses suggested that this intrinsically dynamic interdomain linker forms an additional structural module that plays essential roles in ligand diffusion, recognition, and catalysis. We confirmed that SaCntL stereoselectively carries out the catalysis of D-His but not its enantiomer, L-His, and we found that the N-C linker and active site of SaCntL could accommodate both enantiomers. SaCntL is likely able to bind L-His without catalysis, and as a result, L-His could show inhibitory effects toward SaCntL. These findings provide critical structural and mechanistic insights into CntL, which facilitates a better understanding of the biosynthesis of nicotianamine-like metallophores and the discovery of inhibitors of this process.