Biology subjects

Liang, M.

Publications and source records attributed to Liang, M..

72 records · Page 4Linked to original sources

bioRxiv2021

Antibody Cocktail Exhibits Broad Neutralization against SARS-CoV-2 and SARS-CoV-2 variants

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has precipitated multiple variants resistant to therapeutic antibodies. In this study, 12 high-affinity antibodies were generated from convalescent donors in early outbreaks using immune antibody phage display libraries. Of them, two RBD-binding antibodies (F61 and H121) showed high affinity neutralization against SARS-CoV-2, whereas three S2-target antibodies failed to neutralize SARS-CoV-2. Following structure analysis, F61 identified a linear epitope located in residues G446 -S494, which overlapped with angiotensin-converting enzyme 2 (ACE2) binding sites, while H121 recognized a conformational epitope located on the side face of RBD, outside from ACE2 binding domain. Hence the cocktail of the two antibodies achieved better performance of neutralization to SARS-CoV-2. Importantly, F61 and H121 exhibited efficient neutralizing activity against variants B.1.1.7 and B.1.351, those showed immune escape. Efficient neutralization of F61 and H121 against multiple mutations within RBD revealed a broad neutralizing activity against SARS-CoV-2 variants, which mitigated the risk of viral escape. Our findings defined the basis of therapeutic cocktails of F61 and H121 with broad neutralization and delivered a guideline for the current and future vaccine design, therapeutic antibody development, and antigen diagnosis of SARS-CoV-2 and its novel variants.

immunology↗

bioRxiv2021

Structural insight into host plasma membrane association and assembly of HIV-1 Matrix protein

HIV-1 continues to be a global health concern since AIDS was first recognized by the World Health Organization (WHO). It is estimated that there were 38 million people infected with HIV-1 and 1.5 million deaths in 2019 alone. A better understanding of the details of the HIV late-stage life cycle, involving Pr55Gag attachment to the membrane for the further oligomerization to release virion, will provide us new avenues for potential treatment. Inositol hexakisphosphate (IP6) is an abundant endogenous cyclitol molecule and its binding was linked to the oligomerization of Pr55Gag via the MA domain. However, the binding site of IP6 on MA was unknown and the structural details of this interaction were missing. Here, we present three high-resolution crystal structures of the MA domain in complex with IP6 molecules to reveal its binding mode. Additionally, extensive Differential Scanning Fluorimetry analysis combined with cryo- and ambient-temperature X-ray crystallography and computational biology identify the key residues that participate in IP6 binding. Our data provide novel insights about the multilayered HIV-1 virion assembly process that involves the interplay of IP6 with PIP2, a phosphoinositide essential for the membrane binding of Pr55Gag. IP6 and PIP2 have neighboring alternate binding sites within the same highly basic region (residues 18-33). This indicates that IP6 and PIP2 bindings are not mutually exclusive and may play a key role in coordinating virion particles membrane localization. Based on our three different IP6-MA complex crystal structures, we propose a new model that involves the IP6 coordination of the oligomerization of outer MA and inner CA domain 2D layers during assembly and budding.

microbiology↗

bioRxiv2020

Enteric glial cells respond to a dietary change in the lamina propria in a MyD88-dependent manner

Immune and nervous system sensing are two important ways of detecting inner and outer environmental changes. Immune cell activation in the gut can promote metabolic disorders. However, whether enteric nervous system sensing and activities are also important in metabolic syndromes is not clear. Enteric glial cells (EGCs) are thought to have sensing ability, but little is known about the potential connections between EGC and metabolic disorders. Consuming a modern Western-type high-fat low-fiber diet increases the risk of obesity. Here, we reported that dietary shift from a normal chow diet to a high-fat diet in wild-type (WT) C57BL/6 mice induced a transient emergence of glial fibrillary acidic protein (GFAP)-positive EGC network in the ileal lamina propria, accompanied by an increase of glial-derived neurotrophic factors production. Inhibition of EGC metabolic activity via gliotoxin fluorocitrate or glial-intrinsic deletion of myeloid differentiation factor 88 (Myd88) in mice blocked this dietary change-induced activity. Furthermore, we found a different role of MYD88 in glial cells versus adipocyte in diet-induced obesity. The glial Myd88 knockout mice gained less body weight after HFD feeding compared to the littermate controls. In contrast, adipocyte deletion of Myd88 in mice had no impact on the weight gain but had exacerbated glucose metabolic disorders. Pharmacological interventions of glial activities by fluorocitrate prevented body weight gain in a dietary type- and glial MYD88-independent manner. Collectively, our data reveal a previously unappreciated function of EGC in sensing a dietary shift-induced perturbation and glial activities as a whole may play roles in diet-induced obesity. New & NoteworthyIt is known that obesity and its related metabolic syndrome can damage the neuronal system. However, whether the neuronal system also participates in the development of obesity is unclear. Diet is an important contributing factor to obesity. Our study reveals that consuming a high-fat diet can induce a transient enteric glial cell response via its intrinsic sensing molecule(s). Inhibiting overall glial cell activities may have an impact on the development of the metabolic syndrome.

neuroscience↗

bioRxiv2020

Genomic Resources of Broomcorn Millet: Demonstration and Application of a High-throughput BAC Mapping Pipeline

BackgroundWith high-efficient water-use and drought tolerance, broomcorn millet has emerged as a candidate for food security. To promote its research process for molecular breeding and functional research, a comprehensive genome resource is of great importance. ResultsHerein, we constructed a BAC library for broomcorn millet, generated BAC end sequences based on the clone-array pooled shotgun sequencing strategy and Illumina sequencing technology, and integrated BAC clones into genome by a novel pipeline for BAC end profiling. The BAC library is consisted of 76,023 clones with an average insert length of 123.48 Kb, covering about 9.9-fold of the 850 Mb genome. Of 9,216 clones tested using our pipeline, 8,262 clones were mapped on the broomcorn millet cultivar longmi4 genome. These mapped clones covered 308 of the 829 gaps left by the genome. To our knowledge, this is the only BAC resource for broomcorn millet. ConclusionsWe constructed a high-quality BAC libraray for broomcorn millet and designed a novel pipeline for BAC end profiling. BAC clones can be browsed and obtained from our website (http://eightstarsbio.com/gresource/JBrowse-1.16.5/index.html). The high-quality BAC clones mapped on genome in this study will provide a powerful genomic resource for genome gap filling, complex segment sequencing, FISH, functional research, and genetic engineering of broomcorn millet.

genomics↗

bioRxiv2020

Near-Physiological-Temperature Serial Femtosecond X-ray Crystallography Reveals Novel Conformations of SARS-CoV-2 Main Protease Active Site for Improved Drug Repurposing

The COVID19 pandemic has resulted in 25+ million reported infections and nearly 850.000 deaths. Research to identify effective therapies for COVID19 includes: i) designing a vaccine as future protection; ii) structure-based drug design; and iii) identifying existing drugs to repurpose them as effective and immediate treatments. To assist in drug repurposing and design, we determined two apo structures of Severe Acute Respiratory Syndrome CoronaVirus-2 main protease at ambienttemperature by Serial Femtosecond X-ray crystallography. We employed detailed molecular simulations of selected known main protease inhibitors with the structures and compared binding modes and energies. The combined structural biology and molecular modeling studies not only reveal the dynamics of small molecules targeting main protease but will also provide invaluable opportunities for drug repurposing and structure-based drug design studies against SARS-CoV-2. One Sentence SummaryRadiation-damage-free high-resolution SARS-CoV-2 main protease SFX structures obtained at near-physiological-temperature offer invaluable information for immediate drug-repurposing studies for the treatment of COVID19.

biophysics↗

bioRxiv2020

Common and distinct roles of frontal midline theta and occipital alpha oscillations in coding temporal intervals and spatial distances

Judging how far something is and how long it takes to get there are critical to memory and navigation. Yet, the neural codes for spatial and temporal information remain unclear, particularly the involvement of neural oscillations in maintaining such codes. To address these issues, we designed an immersive virtual reality environment containing teleporters that displace participants to a different location after entry. Upon exiting the teleporters, participants made judgements from two given options regarding either the distance they had travelled (spatial distance condition) or the duration they had spent inside the teleporters (temporal duration condition). We wirelessly recorded scalp EEG while participants navigated in the virtual environment by physically walking on an omnidirectional treadmill and traveling through teleporters. An exploratory analysis revealed significantly higher alpha and beta power for short distance versus long distance traversals, while the contrast also revealed significantly higher frontal midline delta-theta-alpha power, and global beta power increases for short versus long temporal duration teleportation. Analyses of occipital alpha instantaneous frequencies revealed their sensitivity for both spatial distances and temporal durations, suggesting a novel and common mechanism for both spatial and temporal coding. We further examined the resolution of distance and temporal coding by classifying discretized distance bins and 250ms time bins based on multivariate patterns of 2-30 Hz power spectra, finding evidence that oscillations code fine-scale time and distance information. Together, these findings support partially independent coding schemes for spatial and temporal information, suggesting that low-frequency oscillations play important roles in coding both space and time.

neuroscience↗

bioRxiv2020

Simple cloning of large natural product biosynthetic gene cluster by CRISPR/Cas12a-mediated fast direct capturing strategy

Directly cloning of biosynthetic gene clusters (BGCs) from microbial genomes has been revolutionizing the natural product-based drug discovery. However, it is still very challenging to efficiently clone, for example, large (> 80kb) and GC-rich (> 70%), streptomycete originating BGCs. In this study, we developed a simple, fast yet efficient and low-cost in vitro platform for direct cloning large BGCs from streptomycete genomic DNA, named as CAT-FISHING (CRISPR/Cas12a- and Agarose plug-based sysTem for Fast bIoSyntHetIc geNe cluster cloninG), by combining the advantages of CRISPR/Cas12a cleavage and bacterial artificial chromosome (BAC) library construction. CAT-FISHING was demonstrated by directly cloning large DNA fragments ranging from 47 to 139 kb with GC content of > 70% from the S. albus J1074 genome in a relatively efficient manner. Moreover, surugamides, encoded by a captured 87-kb BGC with GC content of 76%, was heterologously expressed in a Streptomyces chassis. These results indicate that CAT-FISHING is a powerful platform for BGCs batch cloning, which would be greatly beneficial to the natural products-based drug discovery. We believe that this system will lead a renaissance of interest in microorganisms as a source for drug development.

synthetic biology↗

bioRxiv2020

Lipidomic profiling of plasma extracellular vesicles as an effective means to evaluate the risk of preterm birth

Preterm birth is the main cause of infant death worldwide and results in a high societal economic burden associated with newborn care. Recent studies have shown that extracellular vesicles play an important role in fetal development during pregnancy. Here, we fully investigated differences in lipids in plasma, microvesicles and exosomes between 27 preterm and 66 full-term pregnant women in the early second trimester (12-24 weeks) using an untargeted lipidomics approach. Independent of other characteristics of samples, we detected 97, 58 and 10 differential features (retention time (RT) and m/z) with identification by multivariate and univariate statistical analyses in plasma, microvesicles and exosomes, respectively. These altered lipids were involved in the formation of the bacterial cell wall and chronic low-level inflammation and oxidative stress. Furthermore, lipids in microvesicles could distinguish patients who experienced preterm labor from controls better than lipids in plasma and exosomes. The candidate lipid biomarkers in microvesicles were also validated by the pseudotargeted lipidomics method. The validation set included 41 preterm and 42 healthy pregnant women. PS (34:0) in microvesicles was able to distinguish preterm birth from healthy pregnancy with higher accuracy. Our study shows that differences in lipids in plasma, microvesicles and exosomes are useful for understanding the underlying mechanisms, early clinical diagnosis and intervention of preterm birth.

biochemistry↗

bioRxiv2020

Genetic diversity and evolution of Hantaan virus in China and its neighbors

BackgroudHantaan virus (HTNV), as one of the pathogenic hantaviruses of HFRS, has raised serious concerns in Eurasia. China and its neighbors, especially Russia and South Korea, are seriously suffered HTNV infections. Recent studies reported genetic diversity and phylogenetic features of HTNV in different parts of China, but the analyses from the holistic perspective are rare. Methodology and Principal FindingsTo better understand HTNV genetic diversity and dynamics, we analyzed all available complete sequences derived from the S and M segments with bio-informatic tools. Our study revealed 11 phylogroups and sequences showed obvious geographic clustering. We found 42 significant amino acid variants sites and 18 of them located in immune epitopes. Nine recombination events and seven reassortment isolates were deteced in our study. Sequences from Guizhou were highly genetic divergent, characterized by the emergence of multiple lineages, recombination and reassortment events. We found that HTNV probably emerged in Zhejiang about 1,000 years ago and the population size expanded from 1980s to 1990s. Bayesian stochastic search variable selection analysis revealed that Heilongjiang, Shaanxi and Guizhou played important roles in HTNV evolution and migration. Conclusions/SignificanceThese findings reveal the original and evolution features of HTNV which might assist in understanding Hantavirus epidemics and would be useful for disease prevention and control. Author summaryHemorrhagic Fever with Renal Syndrome (HFRS) and Hantavirus Pulmonary Syndrome (HPS) are endemic zoonotic infectious diseases caused by hantaviruses that belong to the Family Bunyaviridae. Hantaviruses have gained worldwide attention as etiological agents of emerging zoonotic diseases, with fatality rates ranging from <10% up to 60%. However, our knowledge about the emergence and evolution of HTNV is limited. To get more information about HTNV genetic diversity and phylogenetic features in holistic perspective, we investigated the genetic diversity and spatial distribution of HTNV using all available whole genomic sequences of S and M segments. We also gain insights into the genetic diversity and spatial-temporal dynamics of HTNV. These data can augment traditional approach to infectious disease surveillance and control.

evolutionary biology↗

bioRxiv2019

Ferroptosis contributes to developmental cell death in rice blast

We identified that ferroptosis, an iron-dependent non-apoptotic cell death process, occurs in the rice blast fungus Magnaporthe oryzae, and plays a key role in infection-related development therein. Ferroptosis in the blast fungus was confirmed based on the four basic criteria. We confirmed the dependence of ferroptosis on ferric ions, and optimized C11-BODIPY581/591 as a key sensor for subcellular detection and quantification of lipid peroxides that mediate ferroptotic cell death during the pathogenic growth phase of M. oryzae. In addition, we uncovered an important regulatory function for reduced glutathione and the NADPH oxidases in generating/modulating the superoxide moieties for ferroptotic cell death in Magnaporthe. Ferroptosis was found to be necessary for the specific developmental cell death in conidia during appressorium maturation in rice blast. Such ferroptotic cell death initiated first in the terminal cell and progressed sequentially to the entire conidium. Chelation of iron or chemical inhibition of ferroptosis caused conidial cells to remain viable and led to strong defects in host invasion by M. oryzae. Precocious induction of ferroptosis in a blast-susceptible rice cultivar led to resistance against M. oryzae invasion. Interestingly, ferroptosis and autophagy were found to play inter-reliant or codependent roles in contributing to such precise cell death in M. oryzae conidia during pathogenic differentiation. Our study provides significant molecular insights into understanding the role of developmental cell death and iron homeostasis in infection-associated morphogenesis and in fungus-plant interaction in the blast pathosystem.

cell biology↗

bioRxiv2019

The Golgi Glycoprotein MGAT4D Protects Testicular Germ Cells From Mild Heat Stress

Male germ cells are sensitive to heat stress and testes must be maintained outside the body for optimal fertility. However, no germ cell intrinsic mechanism that protects from heat has been reported. Here, we identify the germ cell specific Golgi glycoprotein MGAT4D as a protector of male germ cells from heat stress. Mgat4d is highly expressed in spermatocytes and spermatids. Unexpectedly, when the Mgat4d gene was inactivated globally or conditionally in spermatogonia, or mis-expressed in spermatogonia, spermatocytes or spermatids, neither spermatogenesis nor fertility were affected. On the other hand, when males were subjected to mild heat stress of the testis (43{degrees}C for 25 min), germ cells with inactivated Mgat4d were markedly more sensitive to the effects of heat stress, and transgenic mice expressing Mgat4d were partially protected from heat stress. Germ cells lacking Mgat4d generally mounted a similar heat shock response to control germ cells, but could not maintain that response. Several pathways activated by heat stress in wild type were induced to a lesser extent in Mgat4d[-/-] heat-stressed germ cells (NF{kappa}B response, TNF and TGF{beta} signaling, Hif1 and Myc genes). Thus, the Golgi glycoprotein MGAT4D is a novel, intrinsic protector of male germ cells from heat stress.

developmental biology↗

bioRxiv2019

Serial femtosecond X-ray diffraction of HIV-1 Gag MA-IP6 microcrystals at ambient temperature

The Human immunodeficiency virus-1 (HIV-1) matrix (MA) domain is involved in the highly regulated assembly process of the virus particles that occur at the host cells plasma membrane. High-resolution structures of the MA domain determined using cryo X- ray crystallography have provided initial insights into the possible steps in the viral assembly process. However, these structural studies have relied on large and frozen crystals in order to reduce radiation damage caused by the intense X-rays. Here, we report the first XFEL study of the HIV-1 MA domains interaction with inositol hexaphosphate (IP6), a phospholipid headgroup mimic. We also describe the purification, characterization and microcrystallization of two MA crystal forms obtained in the presence of IP6. In addition, we describe the capabilities of serial femtosecond X-ray crystallography (SFX) using X-ray free-electron laser (XFEL) to elucidate the diffraction data of MA-IP6 complex microcrystals in liquid suspension at ambient temperature. Two different microcrystal forms of MA-IP6 complex both diffracted to beyond 3.5 [A] resolution, demonstrating the feasibility of using SFX to study the complexes of MA domain of HIV-1 Gag polyprotein with IP6 at near-physiological temperatures. Further optimization of the experimental and data analysis procedures will lead to better understanding of the MA domain of HIV-1 Gag and IP6 interaction at high resolution and provide basis for optimization of the lead compounds for efficient inhibition of the Gag protein recruitment to the plasma membrane prior to virion formation.

biophysics↗

bioRxiv2018

Aminoglycoside ribosome interactions reveal novel conformational states at ambient temperature

The bacterial 30S ribosomal subunit is a primary antibiotic target. Despite decades of discovery, the mechanisms by which antibiotic binding induces ribosomal dysfunction are not fully understood. Ambient temperature crystallographic techniques allow more biologically relevant investigation of how local antibiotic binding site interactions trigger global subunit rearrangements that perturb protein synthesis. Here, the structural effects of 2-deoxystreptamine (paromomycin and sisomicin), a novel sisomicin derivative, N1-methyl sulfonyl sisomicin (N1MS) and the non-deoxystreptamine (streptomycin) aminoglycosides on the ribosome at ambient and cryogenic temperatures were examined. Comparative studies led to three main observations. First, individual aminoglycoside-ribosome interactions in the decoding center were similar for cryogenic vs ambient temperature structures. Second, analysis of a highly conserved GGAA tetraloop of h45 revealed aminoglycoside-specific conformational changes, which are affected by temperature only for N1MS. We report the h44/h45 interface in varying states, that is, engaged, disengaged and in equilibrium. Thirdly, we observe aminoglycoside-induced effects on 30S domain closure, including a novel intermediary closure state, which is also sensitive to temperature. Analysis of three ambient and five cryogenic crystallography datasets reveal a correlation between h44/h45 engagement and domain closure. These observations illustrate the role of ambient temperature crystallography in identifying dynamic mechanisms of ribosomal dysfunction induced by local drug-binding site interactions. Together these data identify tertiary ribosomal structural changes induced by aminoglycoside binding that provides functional insight and targets for drug design.

biophysics↗

bioRxiv2018

The probiotic effectiveness in experimental colitis is correlated with gut microbiome and host genetic features

Current evidence to support extensive use of probiotics in inflammatory bowel disease is limited and factors contribute to the inconsistent effectiveness of clinical probiotic therapy are not completely known. Here, as a proof-of-concept, we utilized Bifidobacterium longum JDM 301, a widely used commercial probiotic strain in China, to study potential factors that may influence the beneficial effect of probiotics in experimental colitis. We found that the probiotic therapeutic effect was varied across individual mouse even with the same genetic background and consuming the same type of food. The different probiotic efficacy was highly correlated with different microbiome features in each mouse. Consumption of a diet rich in fat can change the host sensitivity to mucosal injury-induced colitis but did not change the host responsiveness to probiotic therapy. Finally, the host genetic factor TLR2 was required for a therapeutic effect of B. longum JDM 301. Together, our results suggest that personalized microbiome and genetic features may modify the probiotic therapeutic effect.

microbiology↗

bioRxiv2018

Insufficient fumarase contributes to generating reactive oxygen species in Dahl salt sensitive rats

Dahl SS rats exhibit greater levels of renal medullary oxidative stress and lower levels of fumarase activities. Fumarase insufficiencies can increase reactive oxygen species (ROS), the mechanism of which, however, is not clear. A proteomic analysis indicated fumarase knockdown in HK-2 cells resulted in changes in the expression or activity of NADPH oxidase, mitochondrial respiratory chain complex I and III, ATP synthase subunits, and -oxoglutarate dehydrogenase, all of which are sites of ROS formation. Meantime, the activities of key antioxidant enzymes such as G6PD, 6PGD, GR, GPx and GST increased significantly too. The apparent activation of antioxidant defense appeared insufficient as glutathione precursors, glutathione and GSH/GSSG ratio were decreased. SS rats exhibited changes in redox metabolism similar to HK-2 cells with fumarase knockdown. Supplementation with fumarate and malate, the substrate and product of fumarase, increased and decreased, respectively, blood pressure and the levels of H2O2 and MDA in kidney tissues of SS rats. These results indicate fumarase insufficiencies cause a wide range of changes at several sites of ROS production and antioxidant mechanisms.

molecular biology↗

bioRxiv2018

Structure of the 30S ribosomal decoding complex at ambient temperature

The ribosome translates nucleotide sequences of messenger RNA to proteins through selection of cognate transfer RNA according to the genetic code. To date, structural studies of ribosomal decoding complexes yielding high-resolution data have predominantly relied on experiments performed at cryogenic temperatures. New lightsources like the X-ray free electron laser (XFEL) have enabled data collection from macromolecular crystals at ambient temperature. Here, we report an X-ray crystal structure of the Thermus thermophilus 30S ribosomal subunit decoding complex to 3.45 [A] resolution using data obtained at ambient temperature at the Linac Coherent Light Source (LCLS). We find that this ambient-temperature structure is largely consistent with existing cryogenic-temperature crystal structures, with key residues of the decoding complex exhibiting similar conformations, including adenosine residues 1492 and 1493. Minor variations were observed, namely an alternate conformation of cytosine 1397 near the mRNA channel and the A-site. Our serial crystallography experiment illustrates the amenability of ribosomal microcrystals to routine structural studies at ambient temperature, thus overcoming a long-standing experimental limitation.

biophysics↗

bioRxiv2017

Candidate cancer driver mutations in super-enhancers and long-range chromatin interaction networks

A comprehensive catalogue of the mutations that drive tumorigenesis and progression is essential to understanding tumor biology and developing therapies. Protein-coding driver mutations have been well-characterized by large exome-sequencing studies, however many tumors have no mutations in protein-coding driver genes. Non-coding mutations are thought to explain many of these cases, however few non-coding drivers besides TERT promoter are known. To fill this gap, we analyzed 150,000 cis-regulatory regions in 1,844 whole cancer genomes from the ICGC-TCGA PCAWG project. Using our new method, ActiveDriverWGS, we found 41 frequently mutated regulatory elements (FMREs) enriched in non-coding SNVs and indels (FDR<0.05) characterized by aging-associated mutation signatures and frequent structural variants. Most FMREs are distal from genes, reported here for the first time and also recovered by additional driver discovery methods. FMREs were enriched in super-enhancers, H3K27ac enhancer marks of primary tumors and long-range chromatin interactions, suggesting that the mutations drive cancer by distally controlling gene expression through threedimensional genome organization. In support of this hypothesis, the chromatin interaction network of FMREs and target genes revealed associations of mutations and differential gene expression of known and novel cancer genes (e.g., CNNB1IP1, RCC1), activation of immune response pathways and altered enhancer marks. Thus distal genomic regions may include additional, infrequently mutated drivers that act on target genes via chromatin loops. Our study is an important step towards finding such regulatory regions and deciphering the somatic mutation landscape of the non-coding genome.

cancer biology↗

bioRxiv2017

Authentic Enzyme Intermediates Captured "on-the-fly" by Mix-and-Inject Serial Crystallography

Ever since the first atomic structure of an enzyme was solved, the discovery of the mechanism and dynamics of reactions catalyzed by biomolecules has been the key goal for the understanding of the molecular processes that drive life on earth. Despite a large number of successful methods for trapping reaction intermediates, the direct observation of an ongoing reaction has been possible only in rare and exceptional cases. Here, we demonstrate a general method for capturing enzyme catalysis in-action by mix-and-inject serial crystallography. Specifically, we follow the catalytic reaction of the Mycobacterium tuberculosis -lactamase with the 3rd generation antibiotic ceftriaxone by time-resolved serial femtosecond crystallography. The results reveal, in near atomic detail, antibiotic cleavage and inactivation on the millisecond to second time scales including the crossover from transition state kinetics to steady-state kinetics.\n\nSynopsisAn enzymatically catalyzed reaction is initiated by diffusion based mixing of substrate and followed at runtime by time-resolved serial crystallography using a free electron laser.

biophysics↗