bioRxiv Science⌕ Search

Biology subjects

Ahn, J.

Publications and source records attributed to Ahn, J..

At least 55 records · Page 3Linked to original sources
bioRxiv2022

Mosquito invasion via the global shipping network is slowed in high-risk areas by on-shore and ship-board monitoring

The global shipping network (GSN) has been suggested as a pathway for the establishment and reintroduction of Aedes aegypti and Aedes albopictus primarily via the tire trade. We used historical maritime movement data in combination with an agent-based model to understand invasion risk in the United States Gulf Coast and how the risk of these invasions could be reduced. We found a strong correlation between the total number of cargo ship arrivals at each port and likelihood of arrival by both Ae. aegypti and Ae. albopictus. Additionally, in 2012, 99.2% of the arrivals into target ports had most recently visited ports occupied by both Ae. aegypti and Ae. albopictus, increasing risk of Aedes invasion. Model results indicated that detection and removal of mosquitoes from containers when they are unloaded at a port may be more effective in reducing the establishment of mosquito populations compared to eradication efforts that occur while onboard the vessel, suggesting detection efforts should be focused on unloaded containers. To reduce the risk of invasion and reintroduction of Ae. aegypti and Ae. albopictus, surveillance and control efforts should be employed when containers leave high risk locations and when they arrive in ports at high risk of establishment.

ecology↗
bioRxiv2022

Differential Expression Enrichment Tool (DEET): an interactive atlas of human differential gene expression

Differential gene expression analysis using RNA sequencing (RNA-seq) data is a standard approach for making biological discoveries. Ongoing large-scale efforts to process and normalize publicly available gene expression data enable rapid and systematic reanalysis. While several powerful tools systematically process RNA-seq data, enabling their reanalysis, few resources systematically recompute differentially expressed genes (DEGs) generated from individual studies. We developed a robust differential expression analysis pipeline to recompute 3162 human DEG lists from The Cancer Genome Atlas, Genotype-Tissue Expression Consortium, and 142 studies within the Sequence Read Archive. After measuring the accuracy of the recomputed DEG lists, we built the Differential Expression Enrichment Tool (DEET), which enables users to interact with the recomputed DEG lists. DEET, available through CRAN and RShiny, systematically queries which of the recomputed DEG lists share similar genes, pathways, and TF targets to their own gene lists. DEET identifies relevant studies based on shared results with the users gene lists, aiding in hypothesis generation and data-driven literature review. HighlightsBy curating metadata from uniformly processed human RNA-seq studies, we created a database of 3162 differential expression analyses. These analyses include TCGA, GTEx, and 142 unique studies in SRA, involving 985 distinct experimental conditions. The Differential Expression Enrichment Tool (DEET) allows users to systematically compare their gene lists to this database.

bioinformatics↗
bioRxiv2022

Extracellular heparan 6-O-endosulfatases SULF1 and SULF2 in HNSC and other malignancies

SULF1 and SULF2 are oncogenic in a number of human malignancies, including head and neck squamous cell carcinoma (HNSC). The function of these two heparan sulfate editing enzymes was previously considered largely redundant but the biology of cancer suggests differences that we explore in our RNAseq and RNAScope studies of HNSC and in a pan cancer analysis using the TCGA and CPTAC (proteomics) data. Our studies document a consistent upregulation of SULF1 and SULF2 in HNSC which is associated with poor survival outcomes. SULF2 expression increases in multiple malignancies but less consistently than SULF1, which uniformly increases in the tumor tissues and negatively impacts survival in several types of cancer. Meanwhile, SULF1 showed low expression in cancer cell lines and a scRNAseq study of HNSC shows that SULF1 is not supplied by epithelial tumor cells, like SULF2, but is secreted by cancer associated fibroblasts. Our RNAScope and PDX analysis of the HNSC tissues fully confirm the stromal source of SULF1 and explain the uniform impact of this enzyme on the biology of multiple malignancies. In summary, the SULF1 enzyme, supplied by a subset of cancer associated fibroblasts, is upregulated and negatively impacts HNSC survival at an early stage of the disease progression while the SULF2 enzyme, supplied by tumor cells, impacts survival at later stages of HNSC. This paradigm is common to multiple malignancies and suggests a potential for diagnostic and therapeutic targeting of the heparan sulfatases in cancer diseases.

cancer biology↗
bioRxiv2022

Microbial Risk Score for Capturing Microbial Characteristics, Integrating Multi-omics Data, and Predicting Disease Risk

BackgroundWith the rapid accumulation of microbiome-wide association studies, a great amount of microbiome data are available to study the microbiomes role in human disease and advance the microbiomes potential use for disease prediction. However, the unique features of microbiome data hinder its utility for disease prediction. MethodsMotivated from the polygenic risk score framework, we propose a microbial risk score (MRS) framework to aggregate the complicated microbial profile into a summarized risk score that can be used to measure and predict disease susceptibility. Specifically, the MRS algorithm involves two steps: 1) identifying a sub-community consisting of the signature microbial taxa associated with disease, and 2) integrating the identified microbial taxa into a continuous score. The first step is carried out using the existing sophisticated microbial association tests and pruning and thresholding method in the discovery samples. The second step constructs a community-based MRS by calculating alpha diversity on the identified sub-community in the validation samples. Moreover, we propose a multi-omics data integration method by jointly modeling the proposed MRS and other risk scores constructed from other omics data in disease prediction. ResultsThrough three comprehensive real data analyses using the NYU Langone Health COVID-19 cohort, the gut microbiome health index (GMHI) multi-study cohort, and a large type 1 diabetes cohort separately, we exhibit and evaluate the utility of the proposed MRS framework for disease prediction and multi-omics data integration. In addition, the disease-specific MRSs for colorectal adenoma, colorectal cancer, Crohns disease, and rheumatoid arthritis based on the relative abundances of 5, 6, 12, and 6 microbial taxa respectively are created and validated using the GMHI multi-study cohort. Especially, Crohns disease MRS achieves AUCs of 0.88 ([0.85-0.91]) and 0.86 ([0.78-0.95]) in the discovery and validation cohorts, respectively. ConclusionsThe proposed MRS framework sheds light on the utility of the microbiome data for disease prediction and multi-omics integration, and provides great potential in understanding the microbiomes role in disease diagnosis and prognosis.

microbiology↗
bioRxiv2022

Ingestible capsule for detecting labile inflammatory biomarkers in situ

Transient molecules in the gastrointestinal (GI) tract, such as nitric oxide and hydrogen sulfide, are key signals and mediators of inflammatory bowel disease (IBD). Because these molecules are extremely short-lived in the body, they are difficult to detect. To track these reactive molecules in the GI tract, we have developed a miniaturized device that integrates genetically-engineered probiotic biosensors with a custom-designed photodetector and readout chip. Leveraging the molecular specificity of living sensors, we genetically encoded bacteria to respond to IBD-associated molecules by luminescing. Low-power electronic readout circuits (nanowatt power) integrated into the device convert the light from just 1 L of bacterial culture into a wireless signal. We demonstrate biosensor monitoring in the GI tract of small and large animal models and integration of all components into a sub-1.4 cm3 ingestible form factor capable of supporting wireless communication. The wireless detection of short-lived, disease-associated molecules could support earlier diagnosis of disease than is currently possible, more accurate tracking of disease progression, and more timely communication between patient and their care team supporting remote personalized care.

synthetic biology↗
bioRxiv2021

Using marine cargo traffic to identify countries in Africa with greatest risk of invasion by Anopheles stephensi

Anopheles stephensi is an efficient malaria vector commonly found in South Asia and the Arabian Peninsula, but in recent years it has established as an invasive species in the Horn of Africa (HoA). In this region, An. stephensi was first detected in a livestock quarantine station near a major seaport in Djibouti in 2012, in Ethiopia in 2016, in Sudan in 2018 and Somalia in 2019. Anopheles stephensi often uses artificial containers as larval habitats, which may facilitate introduction through maritime trade as has been seen with other invasive container breeding mosquitoes. If An. stephensi is being introduced through maritime traffic, prioritization exercises are needed to identify locations at greatest risk of An. stephensi introduction for early detection and rapid response, limiting further invasion opportunities. Here, we use UNCTAD maritime trade data to 1) identify coastal African countries which were most highly connected to select An. stephensi endemic countries in 2011, prior to initial detection in Africa, 2) develop a ranked prioritization list of countries based on likelihood of An. stephensi introduction for 2016 and 2020 based on maritime trade alone and maritime trade and habitat suitability, and 3) use network analysis to describe intracontinental maritime trade and eigenvector centrality to determine likely paths of further introduction on the continent if An. stephensi is detected in a new location. Our results show that in 2011, Sudan and Djibouti were ranked as the top two countries with likelihood of An. stephensi introduction based on maritime trade alone, and these were indeed the first two coastal countries in the HoA where An. stephensi was detected. Trade data from 2020 with Djibouti and Sudan included as source populations identify Egypt, Kenya, Mauritius, Tanzania, and Morocco as the top five countries with likelihood of An. stephensi introduction. When factoring in habitat suitability, Egypt, Kenya, Tanzania, Morocco, and Libya are ranked highest. Network analysis revealed that the countries with the highest eigenvector centrality scores, and therefore highest degrees of connectivity with other coastal African nations were South Africa (0.175), Mauritius (0.159), Ghana (0.159), Togo (0.157), and Morocco (0.044) and therefore detection of An. stephensi in any one of these locations has a higher potential to cascade further across the continent via maritime trade than those with lower eigenvector centrality scores. Taken together, these data could serve as tools to prioritize efforts for An. stephensi surveillance and control in Africa. Surveillance in seaports of countries at greatest risk of introduction may serve as an early warning system for the detection of An. stephensi, providing opportunities to limit further introduction and expansion of this invasive malaria vector in Africa.

ecology↗
bioRxiv2021

Targeting properdin - Structure and function of a novel family of tick-derived complement inhibitors

Activation of the serum-resident complement system begins a cascade that leads to activation of membrane-resident complement receptors on immune cells, thus coordinating serum and cellular immune responses. Whilst many molecules act to control inappropriate activation, Properdin is the only known positive regulator of the human complement system. By stabilising the alternative pathway C3 convertase it promotes complement self-amplification and persistent activation boosting the magnitude of the serum complement response by all triggers. We have identified a novel family of alternative pathway complement inhibitors, hereafter termed CirpA. Functional and structural characterisation reveals that CirpA family directly bind to properdin, inhibiting its ability to promote complement activation, and leading to potent inhibition of the complement response in a species specific manner. For the first time this study provides a full functional and structural characterization of a properdin inhibitor, opening avenues for future therapeutic approaches.

immunology↗
bioRxiv2021

Binding affinity prediction for protein-ligand complex using deep attention mechanism based on intermolecular interactions

Accurate prediction of protein-ligand binding affinity is important in that it can lower the overall cost of drug discovery in structure-based drug design. For more accurate prediction, many classical scoring functions and machine learning-based methods have been developed. However, these techniques tend to have limitations, mainly resulting from a lack of sufficient interactions energy terms to describe complex interactions between proteins and ligands. Recent deep-learning techniques show strong potential to solve this problem, but the search for more efficient and appropriate deep-learning architectures and methods to represent protein-ligand complexes continues. In this study, we proposed a deep-neural network for more accurate prediction of protein-ligand complex binding affinity. The proposed model has two important features, descriptor embeddings that contains embedded information about the local structures of a protein-ligand complex and an attention mechanism for highlighting important descriptors to binding affinity prediction. The proposed model showed better performance on most benchmark datasets than existing binding affinity prediction models. Moreover, we confirmed that an attention mechanism was able to capture binding sites in a protein-ligand complex and that it contributed to improvement in predictive performance. Our code is available at https://github.com/Blue1993/BAPA. Author summaryThe initial step in drug discovery is to identify drug candidates for a target protein using a scoring function. Existing scoring functions, however, lack the ability to accurately predict the binding affinity of protein-ligand complexes. In this study, we proposed a deep learning-based approach to extract patterns from the local structures of protein-ligand complexes and to highlight the important local structures via an attention mechanism. The proposed model showed good performance for various benchmark datasets compared to existing models.

bioinformatics↗
bioRxiv2021

Activation of Cytochrome C Peroxidase Function Through Coordinated Foldon Loop Dynamics upon Interaction with Anionic Lipids

Cardiolipin (CL) is a mitochondrial anionic lipid that plays important roles in the regulation and signaling of mitochondrial apoptosis. CL peroxidation catalyzed by the assembly of CL-cytochrome c (cyt c) complexes at the inner mitochondrial membrane is a critical checkpoint. The structural changes in the protein, associated with peroxidase activation by CL and different anionic lipids, are not known at a molecular level. To better understand these peripheral protein-lipid interactions, we compare how phosphatidylglycerol (PG) and CL lipids trigger cyt c peroxidase activation, and correlate functional differences to structural and motional changes in membrane-associated cyt c. Structural and motional studies of the bound protein are enabled by magic angle spinning solid state NMR spectroscopy, while lipid peroxidase activity is assayed by mass spectrometry. PG binding results in a surface-bound state that preserves a nativelike fold, which nonetheless allows for significant peroxidase activity, though at a lower level than binding its native substrate CL. Lipid-specific differences in peroxidase activation are found to correlate to corresponding differences in lipid-induced protein mobility, affecting specific protein segments. The dynamics of omega loops C and D are upregulated by CL binding, in a way that is remarkably controlled by the protein:lipid stoichiometry. In contrast to complete chemical denaturation, membrane-induced protein destabilization reflects a destabilization of select cyt c foldons, while the energetically most stable helices are preserved. Our studies illuminate the interplay of protein and lipid dynamics in the creation of lipid peroxidase-active proteolipid complexes implicated in early stages of mitochondrial apoptosis.

biophysics↗
bioRxiv2020

Identification of Gli1 as a progenitor cell marker for meniscus development and injury repair

Meniscal tears are associated with a high risk of osteoarthritis but currently have no disease-modifying therapies. Using Gli1-CreER tdTomato mice, we found that Gli1+ cells contribute to the development of meniscus horns from 2 weeks of age. In adult mice, Gli1+ cells resided at the superficial layer of meniscus and expressed known mesenchymal progenitor markers. In culture, meniscal Gli1+ cells possessed high progenitor activities under the control of Hh signal. Meniscus injury at the anterior horn induced a quick expansion of Gli1+ cells. Normally, the tissue healed slowly, leading to cartilage degeneration. Ablation of Gli1+ cells further hindered this repair process. Strikingly, intra-articular injection of Gli1+ meniscal cells or an Hh activator right after injury accelerated the bridging of the interrupted ends and attenuated signs of osteoarthritis. Taken together, our work identified a novel progenitor population in meniscus and proposes a new treatment for repairing injured meniscus and preventing osteoarthritis.

developmental biology↗
bioRxiv2020

Separation of coiled-coil structures in lamin A/C is required for elongation of the filament

Intermediate filaments (IFs) commonly have structural elements of a central -helical coiled-coil domain consisting of coil 1a, coil 1b, coil 2, and their flanking linkers. Recently, crystal structure of a long lamin A/C fragment was determined and showed detailed features of a tetrameric unit. The structure further suggested a new binding mode between tetramers, designated eA22, where a parallel overlap of coil 1a and coil 2 is the key interaction. In this study, we investigated the biochemical effects of genetic mutations causing human diseases, focusing on the eA22 interaction. The mutant proteins exhibited either weakened or augmented interactions between coil 1a and coil 2. The ensuing biochemical results indicated that the interaction requires the separation of the coiled-coils in N-terminal of coil 1a and C-terminal of coil 2, coupled with the structural transition in the central -helical rod domain. This study provides insight into the role of coil 1a as a molecular regulator in elongation of the IF proteins.

molecular biology↗
bioRxiv2020

Bone marrow adipogenic lineage precursors (MALPs) promote osteoclastogenesis in bone remodeling and pathologic bone loss

Bone is maintained by coupled activities of bone-forming osteoblasts/osteocytes and bone-resorbing osteoclasts and an alternation of this relationship can lead to pathologic bone loss such as in osteoporosis. It is well known that osteogenic cells support osteoclastogenesis via synthesizing RANKL. Interestingly, our recently identified bone marrow mesenchymal cell population--marrow adipogenic lineage precursors (MALPs) that form a multi-dimensional cell network in bone--was computationally demonstrated to be the most interactive with monocyte-macrophage lineage cells through highly and specifically expressing several osteoclast regulatory factors, including RANKL. Using an adipocyte-specific Adipoq-Cre to label MALPs, we demonstrated that mice with RANKL deficiency in MALPs have a drastic increase of trabecular bone mass in long bones and vertebrae starting from 1 month of age but that their cortical bone is normal. This phenotype was accompanied by diminished osteoclast number and attenuated bone formation at the trabecular bone surface. Reduced RANKL signaling in calvarial MALPs also abolished osteolytic lesions after lipopolysaccharide (LPS) injections. Furthermore, in ovariectomized mice, elevated bone resorption was partially attenuated by RANKL deficiency in MALPs. In summary, our studies identified MALPs as a critical player in controlling bone remodeling during normal bone metabolism and pathological bone loss in a RANKL-dependent fashion.

cell biology↗
bioRxiv2020

Analysis of site and structure specific core fucosylation in liver disease progression using exoglycosidase-assisted data-independent LC-MS/MS

Carbohydrates form one of the major groups of biological macromolecules in living organisms. Many biological processes including protein folding, stability, immune response, and receptor activation are regulated by glycosylation. Fucosylation of proteins regulates such processes and is associated with various diseases including autoimmunity and cancer. Mass spectrometry efficiently identifies structures of fucosylated glycans or sites of core fucosylated N-glycopeptides but quantification of the glycopeptides remains less explored. We performed experiments that facilitate quantitative analysis of the core fucosylation of proteins with partial structural resolution of the glycans and we present results of the mass spectrometric SWATH-type DIA analysis of relative abundances of the core fucosylated glycoforms of 45 glycopeptides derived from 18 serum proteins in liver disease of different etiologies. Our results show that a combination of soft fragmentation with exoglycosidases is efficient at the assignment and quantification of the core fucosylated N-glycoforms at specific sites of protein attachment. In addition, our results show that disease-associated changes in core fucosylation are peptide-dependent and further differ by branching of the core fucosylated glycans. Further studies are needed to verify whether tri- and tetra-antennary core fucosylated glycopeptides could be used as markers of liver disease progression.

biochemistry↗
bioRxiv2020

Cytoplasmic CPSF6 regulates HIV-1 capsid trafficking and infection in a cyclophilin A-dependent manner

Human immunodeficiency virus type 1 (HIV-1) capsid binds host proteins during infection, including cleavage and polyadenylation specificity factor 6 (CPSF6) and cyclophilin A (CypA). We observe that HIV-1 infection induces higher-order CPSF6 formation and capsid-CPSF6 complexes co-traffic on microtubules. CPSF6-capsid complex trafficking is impacted by capsid alterations that reduce CPSF6 binding or by excess cytoplasmic CPSF6 expression, both of which are associated with decreased HIV-1 infection. Higher-order CPSF6 complexes bind and disrupt HIV-1 capsid assemblies in vitro. Disruption of HIV-1 capsid binding to CypA leads to increased CPSF6 binding and altered capsid trafficking, resulting in reduced infectivity. Our data reveal an interplay between CPSF6 and CypA that is important for cytoplasmic capsid trafficking and HIV-1 infection. We propose that CypA prevents HIV-1 capsid from prematurely engaging cytoplasmic CPSF6 and that differences in CypA cellular localization and innate immunity may explain cell-specific variations in HIV-1 capsid trafficking and uncoating. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=131 SRC="FIGDIR/small/136697v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@646829org.highwire.dtl.DTLVardef@1f25cf9org.highwire.dtl.DTLVardef@b5e696org.highwire.dtl.DTLVardef@3593a7_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗
bioRxiv2020

Permeability of the HIV-1 capsid to metabolites modulates viral DNA synthesis

Reverse transcription, an essential event in the HIV-1 lifecycle, requires deoxynucleotide triphosphates (dNTPs) to fuel DNA synthesis, thus requiring penetration of dNTPs into the viral core. The central cavity of the capsid protein (CA) hexamer reveals itself as a plausible channel that allows the passage of dNTPs into assembled capsids. Nevertheless, the molecular mechanism of nucleotide import into the capsid remains unknown. Employing all-atom molecular dynamics simulations, we established that cooperative binding between nucleotides inside a CA hexamer cavity results in energetically-favorable conditions for passive translocation of dNTPs into the HIV-1 capsid. Furthermore, binding of the host cell metabolite inositol hexakisphosphate (IP6) enhances dNTP import, while binding of synthesized molecules like benzenehexacarboxylic acid (BHC) inhibits it. The enhancing effect on reverse transcription by IP6 and the consequences of interactions between CA and nucleotides were corroborated using atomic force microscopy, transmission electron microscopy, and virological assays. Collectively, our results provide an atomistic description of the permeability of the HIV-1 capsid to small molecules and reveal a novel mechanism for the involvement of metabolites in HIV-1 capsid stabilization, nucleotide import and reverse transcription.

biophysics↗
bioRxiv2020

CHLOROPLAST UNUSUAL POSITIONING 1 is a new type of actin nucleation factor in plants

Plants have evolved unique responses to fluctuating light conditions in their environment. One such response, chloroplast photorelocation movement, optimizes photosynthesis under weak light and prevents photodamage under strong light. CHLOROPLAST UNUSUAL POSITIONING 1 (CHUP1) plays a pivotal role in the light-responsive chloroplast movements, which are driven by dynamic reorganization of chloroplast actin (cp-actin) filaments. In this study, we demonstrated that fluorescently tagged CHUP1 colocalized and was coordinately reorganized with cp-actin filaments during chloroplast movement in Arabidopsis thaliana. The resulting asymmetric distribution of CHUP1 was reversibly regulated by the blue light receptor phototropin. X-ray crystallography indicated that the CHUP1 C-terminal domain shares structural similarity with the formin homology 2 (FH2) domain, although there is no sequence similarity between the two domains. The CHUP1 C-terminal domain stimulated actin polymerization in the presence of profilin. We conclude that CHUP1 is a novel, plant-specific actin nucleator that functions in cp-actin-based chloroplast movement. HighlightsO_LIBlue light changes the distribution pattern of CHUP1 C_LIO_LIFormin FH2 and CHUP1 C-terminal domains are structurally similar but not homologous C_LIO_LICHUP1 nucleates and severs actin filaments in vitro C_LIO_LICHUP1 is a novel, plant-specific actin nucleator for chloroplast movement C_LI

plant biology↗
bioRxiv2019

Single cell transcriptomics identifies a unique adipocyte population that regulates bone marrow environment

Bone marrow mesenchymal lineage cells are a heterogeneous cell population involved in bone homeostasis and diseases such as osteoporosis. While it is long postulated that they originate from mesenchymal stem cells (MSCs), the true identity of MSCs and their in vivo bifurcated differentiation routes into osteoblasts and adipocytes remain poorly understood. Here, by employing single cell transcriptome analysis, we identified MSCs and delineated their bi-lineage differentiation paths in young, adult and aging mice. Among several newly discovered mesenchymal subpopulations, one is a distinct population of adipose-lineage cells that we named marrow environment regulating adipose cells (MERAs). MERAs are non-proliferative, post-progenitor cells that express many mature adipocyte markers but are devoid of lipid droplets. They are abundant in the bone marrow of young mice, acting as pericytes and stromal cells that form numerous connections among themselves and with other cells inside bone, including endothelial cells. Genetic ablation of MERAs disrupts marrow vessel structure, promotes de novo bone formation. Taken together, MERAs represent a unique population of adipose lineage cells that exist only in the bone marrow with critical roles in regulating bone and vessel homeostasis.

cell biology↗
bioRxiv2018

Molecular mimicry in deoxy-nucleotide catalysis: the structure of Escherichia coli dGTPase reveals the molecular basis of dGTP selectivity

Deoxynucleotide triphosphate triphosphyohydrolyases (dNTPases) play a critical role in cellular survival and DNA replication through the proper maintenance of cellular dNTP pools by hydrolyzing dNTPs into deoxynucleosides and inorganic triphosphate (PPPi). While the vast majority of these enzymes display broad activity towards canonical dNTPs, exemplified by Sterile Alpha Motif (SAM) and Histidine-aspartate (HD) domain-containing protein 1 (SAMHD1), which blocks reverse transcription of retroviruses in macrophages by maintaining dNTP pools at low levels, Escherichia coli (Ec)-dGTPase is the only known enzyme that specifically hydrolyzes dGTP. However, the mechanism behind dGTP selectivity is unclear. Here we present the free-, ligand (dGTP)- and inhibitor (GTP)-bound structures of hexameric E. coli dGTPase. To obtain these structures, we applied UV-fluorescence microscopy, video analysis and highly automated goniometer-based instrumentation to map and rapidly position individual crystals randomly-located on fixed target holders, resulting in the highest indexing-rates observed for a serial femtosecond crystallography (SFX) experiment. The structure features a highly dynamic active site where conformational changes are coupled to substrate (dGTP), but not inhibitor binding, since GTP locks dGTPase in its apo form. Moreover, despite no sequence homology, dGTPase and SAMHD1 share similar active site and HD motif architectures; however, dGTPase residues at the end of the substrate-binding pocket mimic Watson Crick interactions providing Guanine base specificity, while a 7 [A] cleft separates SAMHD1 residues from dNTP bases, abolishing nucleotide-type discrimination. Furthermore, the structures sheds light into the mechanism by which long distance binding (25 [A]) of single stranded DNA in an allosteric site primes the active site by conformationally \"opening\" a tyrosine gate allowing enhanced substrate binding.\n\nSignificance StatementdNTPases play a critical role in cellular survival through maintenance of cellular dNTP. While dNTPases display activity towards dNTPs, such as SAMHD1 -which blocks reverse transcription of HIV-1 in macrophages- Escherichia coli (Ec)-dGTPase is the only known enzyme that specifically hydrolyzes dGTP. Here we use novel free electron laser data collection to shed light into the mechanisms of (Ec)-dGTPase selectivity. The structure features a dynamic active site where conformational changes are coupled to dGTP binding. Moreover, despite no sequence homology between (Ec)-dGTPase and SAMHD1, both enzymes share similar active site architectures; however, dGTPase residues at the end of the substrate-binding pocket provide dGTP specificity, while a 7 [A] cleft separates SAMHD1 residues from dNTP.

biophysics↗