Biology subjects

Fan, S.

Publications and source records attributed to Fan, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2022

Characterizing the genetic polymorphisms in 370 challenging medically relevant genes using long-read sequencing data from 41 human individuals among 19 global populations

Numerous challenging medically relevant genes (CMRGs) cannot be adequately investigated using next-generation sequencing, hindering the detection of functional variation among these genes. In this study, long-read sequencing data from 41 human individuals across 19 populations were analyzed using the current version of the human reference genome assembly (GRCh38) and a telomere-to-telomere assembly of the human genome (T2T-CHM13). After excluding 142 CMRGs containing windows with a depth of coverage (DoC) significantly deviating from the average DoC value of proteincoding regions in the GRCh38 (138) or T2T-CHM13 (47) assemblies, 179 and 263 CMRGs exhibited copy number variation (CNV) signal in GRCh38 and T2T-CHM13, respectively. In addition, 451 high-impact short variants were detected in 188 CMRGs. Further, some genetic alterations were individual- or continental-superpopulation-specific, suggesting a strong need to consider genetic background differences in future genetic testing and drug design studies. Finally, side-by-side comparisons of short variant calls in CMRGs using NGS and LRS data from 13 samples indicated that 15.79% to 33.96% of high-impact short variants in different individuals could only be detected using LRS data. The results described herein will be an important reference for future clinical and pharmacogenetic studies to further improve precision medicine.

genomics↗

bioRxiv2022

The highly conserved RNA-binding specificity of nucleocapsid protein facilitates the identification of drugs with broad anti-coronavirus activity

The binding of SARS-CoV-2 nucleocapsid (N) protein to both the 5'- and 3'-ends of genomic RNA has different implications arising from its binding to the central region during virion assembly. However, the mechanism underlying selective binding remains unknown. Herein, we performed the high-throughput RNA-SELEX (HTR-SELEX) to determine the RNA-binding specificity of the N proteins of various SARS-CoV-2 variants as well as other {beta}-coronaviruses and showed that N proteins could bind two unrelated sequences, both of which were highly conserved across all variants and species. Interestingly, both these sequence motifs are virtually absent from the human transcriptome; however, they exhibit a highly enriched, mutually complementary distribution in the coronavirus genome, highlighting their varied functions in genome packaging. Our results provide mechanistic insights into viral genome packaging, thereby increasing the feasibility of developing drugs with broad-spectrum anti-coronavirus activity by targeting RNA binding by N proteins.

molecular biology↗

bioRxiv2022

The role of GmXTH1, a new xyloglucan endotransglycosylase/hydrolase from soybean, in regulating soybean root growth at seedling stage

Soybean is an important crop for both food and oil, providing high quality protein and oil for humans and animals. Drought stress has an significant effect on the yield and quality of soybean. In the early vegetative stage of soybean, the developed root system can effectively improve the tolerance of soybean plant to drought. Xyloglucan endoglycosylase/hydrolase (XTH) is a cell wall remodeling enzyme involved in cell wall expansion and degradation. However, there are few studies on XTHs regulation of root growth and response to drought stress in soybean seedlings.The objectives of this study were to clone GmXTH1 gene and determine the effects of over and interference-expression of GmXTH1 on soybean root development at seedling stage. A 1,015 bp full-length of the GmXTH1 gene was cloned from soybean (Glycine max L. Merril) cv. RM18 with a 840 bp open reading frame (ORF) of a putative protein. The over and interference expression vectors of GmXTH1 were transformed into soybean using Agrobacterium method. Transformed strains OEA1, OEA3, IEA4, IEA5 were identified by molecular detection methods. The roots of transformed strains OEA1and OEA3 with GmXTH1 gene overexpression are clearly developed and the root phenotypic attributes, including main root length, primary root number, lateral root length, root surface area, root volume, root dry weight, root fresh weight are higher than the control. In contrast, root attributes of IEA4 and IEA5 with GmXTH1 gene interference expression were significantly lower than those of the control, suggesting that the expression of GmXTH1 gene has an important effect on the soybean root growth and development at seedling stage. Under drought stress conditions, the activities of Superoxide dismutase (SOD), Peroxidase(POD)and Catalase(CAT)in the transgenic strains with an overexpression of the GmXTH1 gene were significantly increased, and the root activity, leaf relative water content and total chlorophyll content were ranked in order of OEAs, CK, and IEAs. In conclusion, GmXHT1 plays an important role in the growth of soybean root at seedling stage. Overexpression of GmXTH1 gene can promote root development of soybean seedling. The results of activities of protective enzymes (SOD, POD, CAT), root activity, leaf relative water content and total chlorophyll content under drought stress indicated that transgenic strains OEA1 and OEA3 with overexpressing GmXTH1 gene were resistant to drought stress, suggesting that GmXTH1 may play a positive role in responding to drought stress at soybean seedling stage.

genetics↗

bioRxiv2022

Nonlinear effect of light intensity on normal axial development of rhesus monkeys

PurposeTo investigate the effects of different indoor lighting intensity (500 lx, 750 lx and 1,000 lx) on normal ocular axial length growth by using juvenal rhesus monkeys. MethodsTwenty-four juvenile monkeys were exposed continuously to normal intensity light (NIL, 500 lx, n=16), medium intensity light (MIL, 750 lx, n=8) and high intensity light (HIL, 1 000 lx, n=8), with a same CCT value (about 3000 K) for 246 days. Axial length, anterior chamber depth, corneal curvature radius were measured at about a monthly interval. ResultsAfter 246 days of light exposure, the growth of axial length of the MIL group (750 lx) were 0.151 {+/-} 0.081 mm and 0.139 {+/-} 0.070 mm in the right and left eyes, respectively, and significantly larger in comparison with the NIL group (500lx, OD: 0.068 {+/-} 0.055 mm, OS: 0.074 {+/-} 0.057 mm) and the HIL group (1000lx,OD:0.063 {+/-} 0.093 mm, OS: 0.084 {+/-} 0.052 mm) monkeys. This effect was stable and robust during the whole experimental period. ConclusionThe effects of different intensity lighting on normal ocular axial development was not linear as most people currently think. We must be cautious when it comes to elevate light intensity in classrooms. Whether this conclusion is correct under lights of other CCT value needs further study.

developmental biology↗

bioRxiv2022

Structure and function of diadenylate cyclase DacM from Mycoplasma ovipneumoniae

Cyclic diadenosine monophosphate (c-di-AMP) is a second-messenger nucleotide that is produced by many bacteria. C-di-AMP can not only regulate bacterial growth, cell-wall homeostasis, ion transport and gene transcription, but can also be recognized by multiple sensor / receptor proteins in infected host cells to trigger an innate immune response. Mycoplasma ovipneumoniae causes non-progressive pneumonia in both sheep and goats. Here, we analyzed c-di-AMP signaling in M. ovipneumoniae, which is a genome-reduced obligately pathogenic bacterium. Our results demonstrate that these bacteria can produce c-di-AMP, and we could identify the diadenylate cyclase, which was named DacM. The enzyme was found to utilize both ATP and ADP to synthesize c-di-AMP, resembling CdaM from a novel family of diadenylate cyclases first found in Mycoplasma pneumoniae. Furthermore, we present the crystal structures of DacM in the apo state and substrate-bound state at 3 [A] and 1.9 [A] resolution, respectively. Mutation of residues Asp112, Gly113, Tyr128, Phe129, and Arg143 surrounding the active sites to Ala were lethal to DacM enzymatic activity. These structures provide valuable insights into the biochemistry of c-di-AMP, and offer a basis for the structure-based design of new drugs for animal husbandry.

pathology↗

bioRxiv2022

Structural and computational insights into the SARS-CoV-2 Omicron RBD-ACE2 interaction

Since SARS-CoV-2 Omicron variant (B.1.1.529) was reported in November 2021, it has quickly spread to many countries and outcompeted the globally dominant Delta variant in several countries. The Omicron variant contains the largest number of mutations to date, with 32 mutations located at spike (S) glycoprotein, which raised great concern for its enhanced viral fitness and immune escape[1-4]. In this study, we reported the crystal structure of the receptor binding domain (RBD) of Omicron variant S glycoprotein bound to human ACE2 at a resolution of 2.6 [A]. Structural comparison, molecular dynamics simulation and binding free energy calculation collectively identified four key mutations (S477N, G496S, Q498R and N501Y) for the enhanced binding of ACE2 by the Omicron RBD compared to the WT RBD. Representative states of the WT and Omicron RBD-ACE2 systems were identified by Markov State Model, which provides a dynamic explanation for the enhanced binding of Omicron RBD. The effects of the mutations in the RBD for antibody recognition were analyzed, especially for the S371L/S373P/S375F substitutions significantly changing the local conformation of the residing loop to deactivate several class IV neutralizing antibodies.

biophysics↗

bioRxiv2021

RFX6-mediated dysregulation defines human β cell dysfunction in early type 2 diabetes

A hallmark of type 2 diabetes (T2D), a major cause of world-wide morbidity and mortality, is dysfunction of insulin-producing pancreatic islet {beta} cells1-3. T2D genome-wide association studies (GWAS) have identified hundreds of signals, mostly in the non-coding genome and overlapping {beta} cell regulatory elements, but translating these into biological mechanisms has been challenging4-6. To identify early disease-driving events, we performed single cell spatial proteomics, sorted cell transcriptomics, and assessed islet physiology on pancreatic tissue from short-duration T2D and control donors. Here, through integrative analyses of these diverse modalities, we show that multiple gene regulatory modules are associated with early-stage T2D {beta} cell-intrinsic defects. One notable example is the transcription factor RFX6, which we show is a highly connected {beta} cell hub gene that is reduced in T2D and governs a gene regulatory network associated with insulin secretion defects and T2D GWAS variants. We validated the critical role of RFX6 in {beta} cells through direct perturbation in primary human islets followed by physiological and single nucleus multiome profiling, which showed reduced dynamic insulin secretion and large-scale changes in the {beta} cell transcriptome and chromatin accessibility landscape. Understanding the molecular mechanisms of complex, systemic diseases necessitates integration of signals from multiple molecules, cells, organs, and individuals and thus we anticipate this approach will be a useful template to identify and validate key regulatory networks and master hub genes for other diseases or traits with GWAS data.

cell biology↗

bioRxiv2021

Severe fever with thrombocytopenia syndrome virus and parthenogenetic Asian longhorned tick Haemaphysalis longicornis (Acari: Ixodidae)

Severe fever with thrombocytopenia syndrome virus (SFTSV) is spreading rapidly in Asia. It is transmitted by Haemaphysalis longicornis (Asian longhorned tick, ALT), which has both parthenogenetically and sexually reproducing populations. Parthenogenetic populations were found in at least 15 provinces in China and strongly correlated with the distribution of SFTS. The distribution of SFTS cases was however poorly correlated with the distribution of populations of bisexual ALTs. Phylogeographic analysis suggested that the parthenogenetic population spread much faster than bisexual population because colonization is independent of sexual reproduction. A higher proportion of parthenogenetic ALTs were collected from migratory birds captured at an SFTS endemic area, implicating the contribution to the long-range movement of ALTs in China. The SFTSV susceptibility of parthenogenetic females was like that of bisexual females under laboratory conditions. These results suggest that parthenogenetic ALT, probably transported by migratory birds, play a major role in the rapid spread of SFTSV. Article Summary LineThe parthenogenetic population of Asian longhorned tick is broadly distributed in China and plays a major role in the long-distance spread of SFTSV and perhaps future invasion of countries outside of Asia.

evolutionary biology↗

bioRxiv2021

Suppression of B-Cell Activation by Human Cord Blood-Derived Stem Cells (CB-SC) through the Galectin-9-Dependent Cell Contact Mechanism

BackgroundWe developed the Stem Cell Educator therapy among multiple clinical trials based on the immune modulations of multipotent cord blood-derived stem cells (CB-SC) on different compartments of immune cells such as T cells and monocytes/macrophages in diabetes and other autoimmune diseases. However, the effects of CB-SC on the B cells remained unclear. To better understand the molecular mechanisms underlying the immune education of CB-SC, we explored the modulations of CB-SC on human B cells. MethodsCB-SC were isolated from human cord blood units and confirmed by flow cytometry with different markers for their purity. B cells were purified by using anti-CD19 immunomagnetic beads from human peripheral blood mononuclear cells (PBMC). Next, the activated B cells were treated in the presence or absence of coculture with CB-SC for 7 days before undergoing flow cytometry analysis of phenotypic change with different markers. RT-PCR was utilized to evaluate the levels of galectin expressions with or without treatment of activated B cells in order to find the key galectin contributing to the B-cell modulation. ResultsFlow cytometry demonstrated that the proliferation of activated B cells was markedly suppressed in the presence of CB-SC, leading to the down-regulation of immunoglobulin productions from the activated B cells. Phenotypic analysis revealed that treatment with CB-SC increased the percentage of IgD+CD27- naive B cells, but decreased the percentage of IgD-CD27+ switched B cells. Transwell assay showed that the immune suppression of CB-SC on B cells was dependent on the manner of cell-cell contact via Gal-9 molecule, as confirmed by the blocking experiment with the anti-Gal-9 monoclonal antibody. Mechanistic studies demonstrated that both calcium levels of cytoplasm and mitochondria were down-regulated after the treatment with CB-SC, causing the decline of mitochondrial membrane potential in the activated B cells. Western blot exhibited that the levels of phosphorylated Akt and Erk1/2 signaling proteins in the activated B cells were also markedly reduced in the presence of CB-SC. ConclusionsCB-SC displayed multiple immune modulations on B cells through the Gal-9-mediated cell-cell contact mechanism and calcium flux/Akt/Erk1/2 signaling pathways. The data advances current understanding about the molecular mechanisms underlying the Stem Cell Educator therapy to treat autoimmune diseases in clinics.

immunology↗

bioRxiv2021

Single-cell foundations of live social gaze interaction in the prefrontal cortex and amygdala

Social gaze interaction powerfully shapes interpersonal communication in humans and other primates. However, little is known about the neural underpinnings of these social behavioral exchanges. Here, we studied neural responses associated with naturalistic, face-to-face, social gaze interactions between pairs of macaques. We examined spiking activity in a large number of neurons spanning four different brain regions involved in social behaviors - the amygdala, orbitofrontal cortex, anterior cingulate cortex, and dorsomedial prefrontal cortex. We observed widespread single-cell representations of social gaze interaction functionalities in these brain regions - social discriminability, social gaze monitoring, and mutual eye contact selectivity. Many of these neurons discriminated looking at social versus non-social stimuli with rich temporal heterogeneity, or parametrically tracked the gaze positions of oneself or the conspecific. Furthermore, many neurons displayed selectivity for mutual eye contact as a function of the initiator or follower of mutual gaze events. Crucially, a significant proportion of neurons coded for more than one of these three signatures of social gaze interaction, supporting the recruitment of partially overlapping neuronal ensembles. Our findings emphasize integrated contributions of the amygdala and prefrontal circuits within the social interaction networks in processing real-life social interactions.

neuroscience↗

bioRxiv2021

Mutation Y453F in the spike protein of SARS-CoV-2 enhances interaction with the mink ACE2 receptor for host adaption

COVID-19 patients transmitted SARS-CoV-2 to minks in the Netherlands in April 2020. Subsequently, the mink-associated virus (miSARS-CoV-2) spilled back over into humans. Genetic sequences of the miSARS-CoV-2 identified a new genetic variant known as "Cluster 5" that contained mutations in the spike protein. However, the functional properties of these "Cluster 5" mutations have not been well established. In this study, we found that the Y453F mutation located in the RBD domain of miSARS-CoV-2 is an adaptive mutation that enhances binding to mink ACE2 and other orthologs of Mustela species without compromising, and even enhancing, its ability to utilize human ACE2 as a receptor for entry. Structural analysis suggested that despite the similarity in the overall binding mode of SARS-CoV-2 RBD to human and mink ACE2, Y34 of mink ACE2 was better suited to interact with a Phe rather than a Tyr at position 453 of the viral RBD due to less steric clash and tighter hydrophobic-driven interaction. Additionally, the Y453F spike exhibited resistance to convalescent serum, posing a risk for vaccine development. Thus, our study suggests that since the initial transmission from humans, SARS-CoV-2 evolved to adapt to the mink host, leading to widespread circulation among minks while still retaining its ability to efficiently utilize human ACE2 for entry, thus allowing for transmission of the miSARS-CoV-2 back into humans. These findings underscore the importance of active surveillance of SARS-CoV-2 evolution in Mustela species and other susceptible hosts in order to prevent future outbreaks.

microbiology↗

bioRxiv2021

The gravitation-driven stress-reduced urothelial barrier in toad bladder urothelium

The urinary bladder urothelial are highly specialized epithelia that protect the underlying tissues from mechanical stress and seal them from the overlying fluid space. To better understand the maintaining permeability induced electrical potential roles played by urothelial in the bladder, we established a protocol of gravitation stress in toad urothelial, observed the transmembrane potential difference variation. MethodThe toad urothelial were mounted in a using chamber which the chamber was separated to two solution spaces, and stable with 0.9% saline solution. The electrodes were settled on the surface of each side of the preparation, serosal side definite as cathode. The using chamber was settled in the centrifugal rotor and under 300 rpm rotation to obtain a vertically +4G gravitation on serosal chamber 5min. Resulta transient transmembrane potential difference increasing was observed after adding CaCl2 (3% solution) in serosal chamber. The amplitude increasing phase included a rapid and a slowly ascending phase. In gravitation stressed urothelial preparation, CaCl2 induced transient phase was significantly increased, furthermore the secondary slowly ascending phase was much more amplified on its amplitude axis and significantly prolonged on the time scale than that evoked in control preparations. The evoked total amplitude increasing were 10 times higher than that in control. ConclusionThe urinary bladder epithelial layer has a structure which regulates ion permeability as a barrier. The tight junction plays an important role as the intercellular coupling in the apical side of the epithelial cell. On the other hand, it is known that the ion channel exists on the epithelial cell membrane and regulates the physiological process. The gravitation stress weakened the tight junction. The transmembrane potential difference was enhanced both on its amplitude and prolonged time. The gravitation stress induced hyperpolarization that evoked by CaCl2 is one kind of Cl- transfer from serosal chamber in which high Ca2+ in the urothelial basal membrane activated the calcium-activated chloride channels. This outwardly rectifying chloride channel induced hyperpolarization can be blocked by Nppb.

physiology↗

bioRxiv2021

Ras guanine nucleotide exchange factor RasGRP1 promotes acute inflammatory response and restricts inflammation-contributed cancer cell growth

Acute inflammatory response needs to be tightly regulated for promoting the elimination of pathogens and preveting the risk of tumorigenesis, but the mechanism has not been fully elucidated. Here, we report that Ras guanine nucleotide releasing protein 1 (RasGRP1) plays a bifunctional regulator that promotes acute inflammation and inhibits inflammation-associated cancer. At the mRNA level, RasGRP1 strengthens the inflammatory response by functioning as a competing endogenous RNA to specifically promote IL-6 expression by sponging let-7a. In vivo overexpression of the RasGRP1 3 untranslated region significantly aggravated lipopolysaccharide-induced systemic inflammation and dextran sulphate sodium-induced colitis in IL-6+/+ mice but not in IL-6-/- mice. At the protein level, RasGRP1 restricts the growth of inflammation-contributed cancer cells by impairing EGFR-SOS1-Ras-AKT signalling. Tumour patients with high RasGRP1 expression showed a better clinical outcome than those with low expression. Considering acute inflammation rarely leads to tumorigenesis, this work reveals that RasGRP1 is an essential bifunctional regulator for acute inflammatory response.

immunology↗

bioRxiv2021

The global carrier frequency and genetic prevalence of Upshaw-Schulman syndrome

PurposeUpshaw-Schulman syndrome (USS) is an autosomal recessive disease of thrombotic microangiopathy, caused by pathogenic variants in ADAMTS13. We aimed to (1) perform data mining pathogenicity of ADAMTS13 variants, (2) estimate carrier frequency and genetic prevalence of USS from gnomAD data, and (3) curated ADAMTS13 gene pathogenic variants dataset. MethodsPubMed and Scopus were comprehensive retrieved. All previously reported pathogenic ADAMTS13 variants were compiled and annotated with gnomAD allele frequencies. Pooled global and population-specific carrier frequency and genetic prevalence for USS were calculated using Hardy-Weinberg equation. Resultswe mined reported disease-causing variants, of these were present in gnomAD exomes v2.1.1, filtering by allele frequency, pathogenicity of variants were classified by American College of Medical Genetics and Genomics criteria. The genetic prevalence and carrier frequency of USS was 0.43 per 1 million (95% CI: [0.36, 0.55]) and 1.31 per thousand, respectively. Combining known with novel pathogenic/likely pathogenic variants, the genetic prevalence and carrier frequency are 1.1 per 1 million (95% CI: [0.89, 1.37]) and 2.1 per thousand, respectively. Conclusionthe genetic prevalence and carrier frequency of Upshaw-Schulman syndrome are within range of previously rough estimated.

bioinformatics↗

bioRxiv2021

Methylation of KRAS at lysine 182 and 184 by SETD7 promotes KRAS degradation

Oncogenic KRAS mutations are considered to be a key driver for initiation and progression in non-small-cell lung cancer (NSCLC). However, how post-translational modifications (PTMs) of KRAS, especially methylation, modify KRAS activity and downstream signals remain largely unclear. Here, we showed that SET domain containing histone lysine methyltransferase 7 (SETD7) interacts with KRAS and methylates KRAS at lysine 182 and 184. SETD7-mediated methylation of KRAS led to degradation of KRAS and attenuation of the RAS/MEK/ERK signaling cascade, endowing SETD7 with a potent tumor-suppressive role in NSCLC, both in vitro and in vivo. Mechanistically, RABGEF1, a ubiquitin E3 ligase of KRAS, was recruited and promoted KRAS degradation in a K182/K184 methylation-dependent manner. Notably, low SETD7 or RABGEF1 expression was associated with poor prognosis in lung adenocarcinoma patients. Taken together, our results establish a novel connection between lysine methylation and KRAS protein stability, in addition to elucidating a tumor-suppressive function of SETD7 that operates via modulation of oncogenic RAS signaling.

cancer biology↗

bioRxiv2020

Zinc Dependent Conformational Changes in the Cation Diffusion Facilitator YiiP from S. oneidensis

YiiP is a secondary transporter that couples Zn2+ transport to the proton motive force. Structural studies of YiiP from prokaryotes as well as Znt8 from humans revealed three different Zn2+ sites and a conserved homodimeric architecture. These structures define the inward-facing and outward-facing states that characterize the archetypal alternating access mechanism of transport. To study effects of Zn2+ binding on the conformational transition, we have used a YiiP/Fab complex for single-particle cryo-EM together with Molecular Dynamics simulation to compare structures of YiiP from S. oneidensis in presence and absence of Zn2+. Without Zn2+, YiiP exhibits enhanced flexibility and adopts a novel conformation that appears to be an intermediate state. The transition closes a hydrophobic gate and is controlled by the Zn2+ site at the cytoplasmic membrane interface. This work enhances our understanding of individual Zn2+ binding sites and their role in the conformational dynamics that governs the transport cycle.

biophysics↗

bioRxiv2020

Dextran sulfate sodium salt corrupted colonic crypts declined the smooth muscle tension in mouse large intestine

Ulcerative colitis is one kind of colonic mucosa damage, shows high number of inflammatory epithelial cells. Dextran sulfate sodium salt (DSS) induce a milder onset of colitis or a more aggressive response. It may damage the protective effects on intestinal barrier. In this study, we investigated the damaging of colon crypts, evaluated the smooth muscle tension beneath corrupted crypts in DSS exposed mice. Methodsfemale specific-pathogen-free BALB/C mice (n=16) are randomly divided as: group A: control mice (n=4); group B: DSS-mice (colitis, 5% DSS in drink water, days 1 to 7, n = 12). The DSS is replaced every 2 days. On day 8, mice colons are excised from the colon-cecal junction to the anus. The distal colon segment is longitude incision and aberrant crypt area are determined by methylene blue staining method. The smooth muscle strip is separated and prepared for passive tension tests. The rest segment is fixed with 10% formalin and embedded in paraffin. Histological scores are evaluated in hematoxylin-eosin staining section: crypt damage (none = 0, basal 1/3 damaged = 1, basal 2/3 damaged = 2, only the surface epithelium is intact = 3, and entire crypt and epithelium are lost = 4). The smooth muscle passive tension beneath the aberrant crypt area in DSS-mice are tested and compared with the preparations from control mice. ResultsIn DSS uptake mice, the inflammation in large intestine mucosa damaged crypts with architectural distortions on day 7 (n=7). In crypts damage area, the smooth muscle passive tension and relative myogenic spontaneous contraction parameters are significantly reduced under the high preload conditions. The maximum rate of change of velocity of spontaneous contraction was noticeable attenuated. ConclusionOur findings demonstrate that low dosage DSS water drink result in corrupted colonic crypts. The corrupted crypts damage the large intestinal epithelium barrier, affect the smooth muscle functions, which declined in myogenic spontaneous contraction under the preload. This further may reduce the peristalsis in large intestine.

physiology↗

bioRxiv2020

Re-discovery of PF-3845 as a new chemical scaffold inhibiting phenylalanyl-tRNA synthetase in Mycobacterium tuberculosis

Mycobacteria tuberculosis (Mtb) remains the deadliest pathogenic bacteria worldwide. The search for new antibiotics to treat drug-sensitive as well as drug-resistant tuberculosis has become a priority. The essential enzyme phenylalanyl-tRNA synthetase (PheRS) is an antibacterial drug target because of the large differences between bacterial and human PheRS counterparts. In a high-throughput screening of 2148 bioactive compounds, PF-3845, which is a known inhibitor of human fatty acid amide hydrolase (FAAH), was identified inhibiting Mtb PheRS at Ki [~]0.73 {+/-} 0.055 {micro}M. The inhibition mechanism was studied with enzyme kinetics, protein structural modelling and crystallography, in comparison to a PheRS inhibitor of the noted phenyl-thiazolylurea-sulfonamide class. The 2.3-[A] crystal structure of Mtb PheRS in complex with PF-3845 revealed its novel binding mode, in which a trifluoromethyl-pyridinylphenyl group occupies the Phe pocket while a piperidine-piperazine urea group binds into the ATP pocket through an interaction network enforced by a sulfate ion. It represents the first non-nucleoside bi-substrate competitive inhibitor of bacterial PheRS. PF-3845 inhibits the in vitro growth of Mtb H37Rv at [~]24 {micro}M, and the potency of PF-3845 increased against Mtb pheS-FDAS, suggesting on target activity in mycobacterial whole cells. PF-3845 does not inhibit human cytoplasmic or mitochondrial PheRSs in biochemical assay, which can be explained from the crystal structures. Further elaboration of the piperidine-piperazine urea moiety by medicinal chemistry effort will produce potential antibacterial lead with improved selectivity on the cellular level.

biochemistry↗