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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2015

Hypothetical protein CT 398 ( C ds Z ) interacts with σ 54 ( R po N )‐holoenzyme and the type III secretion export apparatus in Chlamydia trachomatis

Abstract A significant challenge to bacteriology is the relatively large proportion of proteins that lack sufficient sequence similarity to support functional annotation (i.e. hypothetical proteins). The aim of this study was to apply protein structural homology to gain insights into a candidate protein of unknown function (CT398) within the medically important, obligate intracellular bacterium Chlamydia trachomatis . C. trachomatis is a major human pathogen responsible for numerous infections throughout the world that can lead to blindness and infertility. A 2.12 Å crystal structure of hypothetical protein CT398 was determined that was comprised of N‐terminal coiled‐coil and C‐terminal Zn‐ribbon domains. The structure of CT398 displayed a high degree of structural similarity to FlgZ ( Fl a g ellar‐associated z inc‐ribbon domain protein) from Helicobacter pylori . This observation directed analyses of candidate protein partners of CT398, revealing interactions with two paralogous type III secretion system (T3SS) ATPase‐regulators (CdsL and FliH) and the alternative sigma factor RpoN (σ 54 ). Furthermore, genetic introduction of a conditional expression, affinity‐tagged construct into C. trachomatis enabled the purification of a CT398‐RpoN‐holoenzyme complex, suggesting a potential role for CT398 in modulating transcriptional activity during infection. The interactions reported here, in tandem with previous FlgZ studies in H. pylori , indicate that CT398 functions as a regulator of several key areas of chlamydial biology throughout the developmental cycle. Accordingly, we propose that CT398 be named CdsZ ( C ontact‐ d ependent s ecretion‐associated z inc‐ribbon domain protein).

Barta, Michael L.↗
DOE OSTI2018

Screening and identification of potential protein biomarkers for evaluating the efficacy of intensive therapy in pulmonary tuberculosis

Highlights: • The one-year cure rate was significantly higher in sputum negative patients after 2-months intensive therapy. • Four potential biomarkers were found for evaluating the efficacy of 2-months intensive therapy in TB patients. • CO7 and ANGT could serve as potential biomarkers to evaluate the bacteriological condition of Mycobacterium tuberculosis after intensive therapy. This research aimed to discover potential biomarkers for evaluating the therapeutic efficacy of intensive therapy in pulmonary tuberculosis (TB). Protein profiles in 2-months intensively treated TB patients, untreated TB patients, and healthy controls were investigated with iTRAQ-2DLC-MS/MS technique. 71 differential proteins were identified in 2-months intensively treated TB patients. Significant differences in complement component C7 (CO7), apolipoprotein A-IV (APOA4), apolipoprotein C-II (APOC2), and angiotensinogen (ANGT) were found by ELISA validation. CO7 and ANGT were also found significantly different in sputum negative patients, compared with sputum positive patients after intensive treatment. Clinical analysis showed that after 2-months intensive treatment several indicators were significantly changed, and the one-year cure rate of sputum negative patients were significantly higher than sputum positive patients. Diagnostic models consisting of APOC2, CO7 and APOA4 were established to distinguish intensively treated TB patients from untreated TB patients and healthy controls with the AUC value of 0.910 and 0.935. Meanwhile, ANGT and CO7 were combined to identify sputum negative and sputum positive TB patients after intensive treatment with 89.36% sensitivity, 71.43% specificity, and the AUC value of 0.853. The results showed that APOC2, CO7, APOA4, and ANGT may be potential biomarkers for evaluating the efficacy of intensive anti-TB therapy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2019

Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging

The agar culture plate has played a crucial role in bacteriology since the origins of the discipline. However, plating, inoculating, and waiting for microbes to develop colonies that are visible is time-consuming. In this work, we explored if white-light interferometry (WLI) would be a practical tool for reducing the wait time required before colonies can be identified and counted. High resolution surface profile imaging was used for non-destructive characterization and counting of bacterial colonies on agar before they became visible to the naked eye. The three-dimensional (3D) morphology of Gram-negative ( Pseudomonas fluorescens ) and Gram-positive ( Bacillus thuringiensis ) bacterial species were visualized over time by collecting surface profiles of colonies on agar plates with high vertical resolution (3-5 nanometers) and large field of view (3-5 mm). This unique combination of sensitive vertical resolution at low magnification enabled simultaneous detection of hundreds of microcolonies. The colonies were accurately counted within the first few hours after plating with results comparing favorably to counts made by traditional methods. Since the imaging is non-destructive it was used to track single cells multiplying into small colonies and the volumetric changes in the colony were used to accurately measure colony growth rates over time. Based on the results herein, bioimaging with WLI was demonstrated as a powerful bacterial culture assay. Fast non-destructive counting of colony-forming units in a culture and simultaneous measurement of bacterial growth rates with this method suggest that it may lead to beneficial future applications in research, clinical, and other settings.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Formation of Pseudomonas aeruginosa Biofilms in Full-thickness Scald Burn Wounds in Rats

Using Sprague-Dawley rats (350–450 g; n = 61) and the recently updated Walker-Mason rat scald burn model, we demonstrated that Pseudomonas aeruginosa readily formed biofilms within full-thickness burn wounds. Following the burn, wounds were surface-inoculated with P. aeruginosa in phosphate-buffered saline (PBS), while sterile PBS was used for controls. On post-burn days 1, 3, 7, and 11, animals were euthanized and samples collected for quantitative bacteriology, bacterial gene expression, complete blood cell counts, histology, and myeloperoxidase activity. Robust biofilm infections developed in the full-thickness burn wounds inoculated with 1 × 10 4 CFU of P. aeruginosa . Both histology and scanning electron microscopy showed the pathogen throughout the histologic cross-sections of burned skin. Quantigene analysis revealed significant upregulation of alginate and pellicle biofilm matrix genes of P. aeruginosa within the burn eschar. Additionally, expression of P. aeruginosa proteases and siderophores increased significantly in the burn wound environment. Interestingly, the host’s neutrophil response to the pathogen was not elevated in either the eschar or circulating blood when compared to the control burn. This new full-thickness burn biofilm infection model will be used to test new anti-biofilm therapies that may be deployed with soldiers in combat for immediate use at the site of burn injury on the battlefield.

Science & Technology - Other Topics↗
DOE OSTI2015

International code of nomenclature of prokaryotes

Here, this volume contains the edition of the International Code of Nomenclature of Prokaryotes that was presented in draft form and available for comment at the Plenary Session of the Fourteenth International Congress of Bacteriology and Applied Microbiology (BAM), Montréal, 2014, together with updated lists of conserved and rejected bacterial names and of Opinions issued by the Judicial Commission. As in the past it brings together those changes accepted, published and documented by the ICSP and the Judicial Commission since the last revision was published. Several new appendices have been added to this edition. Appendix 11 addresses the appropriate application of the Candidatus concept, Appendix 12 contains the history of the van Niel Prize, and Appendix 13 contains the summaries of Congresses.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Phylogeography of the blacklegged tick ( Ixodes scapularis ) throughout the USA identifies candidate loci for differences in vectorial capacity

Abstract The blacklegged tick ( Ixodes scapularis ( Journal of the Academy of Natural Sciences of Philadelphia , 1821, 2 , 59)) is a vector of Borrelia burgdorferi sensu stricto ( s.s .) ( International Journal of Systematic Bacteriology , 1984, 34 , 496), the causative bacterial agent of Lyme disease, part of a slow‐moving epidemic of Lyme borreliosis spreading across the northern hemisphere. Well‐known geographical differences in the vectorial capacity of these ticks are associated with genetic variation. Despite the need for detailed genetic information in this disease system, previous phylogeographical studies of these ticks have been restricted to relatively few populations or few genetic loci. Here we present the most comprehensive phylogeographical study of genome‐wide markers in I. scapularis , conducted by using 3RAD (triple‐enzyme restriction‐site associated sequencing) and surveying 353 ticks from 33 counties throughout the species' range. We found limited genetic variation among populations from the Northeast and Upper Midwest, where Lyme disease is most common, and higher genetic variation among populations from the South. We identify five spatially associated genetic clusters of I. scapularis . In regions where Lyme disease is increasing in frequency, the I. scapularis populations genetically group with ticks from historically highly Lyme‐endemic regions. Finally, we identify 10 variable DNA sites that contribute the most to population differentiation. These variable sites cluster on one of the chromosome‐scale scaffolds for I. scapularis and are within identified genes. Our findings illuminate the need for additional research to identify loci causing variation in the vectorial capacity of I. scapularis and where additional tick sampling would be most valuable to further understand disease trends caused by pathogens transmitted by I. scapularis .

3RAD↗
DOE OSTI2024

Detection of anaerobic and aerobic bacteria from commercial tattoo and permanent makeup inks

ABSTRACT Tattooing and use of permanent makeup (PMU) have dramatically increased over the last decade, with a concomitant increase in ink-related infections. Studies have shown evidence that commercial tattoo and PMU inks are frequently contaminated with pathogenic microorganisms. Considering that tattoo inks are placed into the dermal layer of the skin where anaerobic bacteria can thrive and cause infections in low-oxygen environments, the prevalence of anaerobic and aerobic bacteria should be assessed in tattoo and PMU inks. In this study, we tested 75 tattoo and PMU inks using the analytical methods described in the FDA Bacteriological Analytical Manual Chapter 23 for the detection of both aerobic and anaerobic bacterial contamination, followed by 16S rRNA gene sequencing for microbial identification. Of 75 ink samples, we found 26 contaminated samples with 34 bacterial isolates taxonomically classified into 14 genera and 22 species. Among the 34 bacterial isolates, 19 were identified as possibly pathogenic bacterial strains. Two species, namelyCutibacterium acnes(four strains) andStaphylococcus epidermidis(two strains) were isolated under anaerobic conditions. Two possibly pathogenic bacterial strains,Staphylococcus saprophyticusandC. acnes, were isolated together from the same ink samples (n= 2), indicating that tattoo and PMU inks can contain both aerobic (S. saprophyticus) and anaerobic bacteria (C. acnes). No significant association was found between sterility claims on the ink label and the absence of bacterial contamination. The results indicate that tattoo and PMU inks can also contain anaerobic bacteria. IMPORTANCE The rising popularity of tattooing and permanent makeup (PMU) has led to increased reports of ink-related infections. This study is the first to investigate the presence of both aerobic and anaerobic bacteria in commercial tattoo and PMU inks under aerobic and anaerobic conditions. Our findings reveal that unopened and sealed tattoo inks can harbor anaerobic bacteria, known to thrive in low-oxygen environments, such as the dermal layer of the skin, alongside aerobic bacteria. This suggests that contaminated tattoo inks could be a source of infection from both types of bacteria. The results emphasize the importance of monitoring these products for both aerobic and anaerobic bacteria, including possibly pathogenic microorganisms.

Biotechnology & Applied Microbiology↗
DOE OSTI2014

The Salmonella type III secretion system virulence effector forms a new hexameric chaperone assembly for export of effector/chaperone complexes

Bacteria hijack eukaryotic cells by injecting virulence effectors into host cytosol with a type III secretion system (T3SS). Effectors are targeted with their cognate chaperones to hexameric T3SS ATPase at the bacterial membrane's cytosolic face. In this issue of the Journal of Bacteriology , Roblin et al. (P. Roblin, F. Dewitte, V. Villeret, E. G. Biondi, and C. Bompard, J Bacteriol 197:688–698, 2015, http://dx.doi.org/10.1128/JB.02294-14) show that the T3SS chaperone SigE of Salmonella can form hexameric rings rather than dimers when bound to its cognate effector, SopB, implying a novel multimeric association for chaperone/effector complexes with their ATPase.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Machine Learning of Bacterial Transcriptomes Reveals Responses Underlying Differential Antibiotic Susceptibility

In vitro antibiotic susceptibility testing often fails to accurately predict in vivo drug efficacies, in part due to differences in the molecular composition between standardized bacteriologic media and physiological environments within the body. Here, we investigate the interrelationship between antibiotic susceptibility and medium composition in Escherichia coli K-12 MG1655 as contextualized through machine learning of transcriptomics data. Application of independent component analysis, a signal separation algorithm, shows that complex phenotypic changes induced by environmental conditions or antibiotic treatment are directly traced to the action of a few key transcriptional regulators, including RpoS, Fur, and Fnr. Integrating machine learning results with biochemical knowledge of transcription factor activation reveals medium-dependent shifts in respiration and iron availability that drive differential antibiotic susceptibility. By extension, the data generation and data analytics workflow used here can interrogate the regulatory state of a pathogen under any measured condition and can be applied to any strain or organism for which sufficient transcriptomics data are available.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Cilantro microbiome before and after nonselective pre-enrichment for Salmonella using 16S rRNA and metagenomic sequencing

Background: Salmonella enterica is a common cause of foodborne gastroenteritis in the United States and is associated with outbreaks in fresh produce such as cilantro. Salmonella culture-based detection methods are complex and time consuming, and improvements to increase detection sensitivity will benefit consumers. In this study, we used 16S rRNA sequencing to determine the microbiome of cilantro. We also investigated changes to the microbial community prior to and after a 24-hour nonselective pre-enrichment culture step commonly used by laboratory analysts to resuscitate microorganisms in foods suspected of contamination with pathogens. Cilantro samples were processed for Salmonella detection according to the method in the United States Food and Drug Administration Bacteriological Analytical Manual. Genomic DNA was extracted from culture supernatants prior to and after a 24-hour nonselective pre-enrichment step and 454 pyrosequencing was performed on 16S rRNA amplicon libraries. A database of Enterobacteriaceae 16S rRNA sequences was created, and used to screen the libraries for Salmonella, as some samples were known to be culture positive. Additionally, culture positive cilantro samples were examined for the presence of Salmonella using shotgun metagenomics on the Illumina MiSeq. Results: Time zero uncultured samples had an abundance of Proteobacteria while the 24-hour enriched samples were composed mostly of Gram-positive Firmicutes. Shotgun metagenomic sequencing of Salmonella culture positive cilantro samples revealed variable degrees of Salmonella contamination among the sequenced samples. Conclusions: Our cilantro study demonstrates the use of high-throughput sequencing to reveal the microbiome of cilantro, and how the microbiome changes during the culture-based protocols employed by food safety laboratories to detect foodborne pathogens. Finding that culturing the cilantro shifts the microbiome to a predominance of Firmicutes suggests that changing our culture-based methods will improve detection sensitivity for foodborne enteric pathogens

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Radiochemically-supported microbial communities. A potential mechanism for biocolloid production of importance to actinide transport

The work described here revealed the presence of diverse microbial communities located across 19 subsurface sites at the NNSS/NTTR and nearby locations. Overall, the diversity of microorganisms was high for subsurface habitats and variable between sites. As of this writing, preparations are being made to combine the Illumina sequences and 16S rRNA clone libraries with other non-NNSS/NTTR well sites of Southern Nevada Regional Flow System for a publication manuscript describing our very broad landscape scale survey of subsurface microbial diversity. Isolates DRI-13 and DRI-14 remain to be fully characterized and named in accordance with the conventions established by Bergey's Manual of Systematic Bacteriology. In preparation to be published, these microorganisms will be submitted to the American Type Culture Collection (ATCC) and the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ).It is anticipated that the data resulting from this study in combination with other data sets that will allow us to produce a number of publications that will be impactful to the subsurface microbiology community.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2018

Evaluation of the Impacts of Heat Exchanger Operation on Quality of Water Used as Heat Source and Sink

To help inform future decisions about the subsequent uses of water that has served as a heat source and sink in the heat exchanger component of a geothermal heat pump system, ORNL performed an independent assessment of data characterizing the quality of water that had passed through a heat exchanger supporting a geothermal heat pump in a school building. Water supply to the heat exchanger is treated drinking water from a public water supply system that derives its water from a groundwater source. Water analyses and bacteriological measurements collected from heat exchanger and inflow and outflow lines were supplied to ORNL for review. Data were compared with regulatory standards and inflow and outflow data were compared to identify changes occurring in the water resulting from its passage through the heat exchanger. Review of the data identified no conditions that would prevent the use of heat exchange outflow water for water supply. Water quality conforms with applicable regulatory standards. There were no discernible differences between inflow and outflow water quality for any parameters other than water temperature and formation of heterotrophic bacterial biofilms. Changes in water temperature are an expected result of the operation of the heat exchanger system and do not by themselves affect the suitability of the water for human consumption. Bacterial biofilm formation can be affected by water temperature, but the data did not show a consistent or statistically significant relationship between temperature parameters and biofilm formation, and the observations of biofilms formed in the heat exchanger water lines do not affect the suitability of the outflow water for water supply. Legionella, a bacterial genus that has been associated with outbreaks of waterborne pathogenic disease, was detected in one biofilm sample, and the water company is performing follow-up testing to assess the significance of this detection.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Recalls of tattoo and permanent makeup inks in the United States and a follow-up microbiological survey of inks with a previous recall history

In this study, we collected voluntary recall records of tattoo and permanent makeup ink from the U.S. Food and Drug Administration (US FDA) Enforcement Report Database. The recall records contain information, such as recall date, manufacturer, ink color, reason for recall, and the microorganisms detected from the ink samples. Between 2003 and 2021, a total of 15 voluntary tattoo ink recalls occurred in the U.S. market, involving over 200 tattoo inks marketed by 13 manufacturers and one distributor. Fourteen recalls were due to microbial contamination, and one recall was due to allergic reaction. As follow-up, a microbiological survey of 28 tattoo inks of new batches from seven manufacturers having products that were previously recalled was conducted. Aerobic plate count (APC) and enrichment culture methods based on the FDA’s Bacteriological Analytical Manual (BAM) were used to detect microbial contamination. The results revealed that six out of 28 tattoo inks were contaminated with bacteria and were produced by two manufacturers. The level of microbial contamination was less than 250 CFU/g in three of the tattoo inks and between 1 × 10 3 and 1 × 10 5 CFU/g in the other three inks. Eleven bacterial isolates were identified, including spore-forming Bacillus -related species and potentially pathogenic species. Overall, this study shows that some tattoo ink products produced by manufacturers with a recall history continue to be contaminated with microorganisms. This highlights the need for ongoing monitoring and quality control of such products.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Editorial: Water supply sustainability and challenges in Asian megadeltas under global change

Globally, riverine deltas comprise barely 1% of land area, but they support the livelihoods of more than 500 million people (Moorhouse et al., 2021). The Asian megadeltas (e.g., Indus, Bengal, Mekong, Red River, and Yangtze) and their riverine floodplains are the most dynamic economic and environmental systems in the world. These low-lying coastal landforms support surface water bodies and highly productive aquifer systems. However, water resources of the Asian megadeltas are extremely vulnerable to geogenic (e.g., arsenic) and anthropogenic (e.g., bacteria) contamination, increased water salinisation from rising sea levels (Figure 1), and chemicals (e.g., fertilizers, pesticides) from land-use practices (Mukherjee et al., 2024) – all of which are threatening the sustainability of water supply and food security in the region. This Research Topic collection features five original articles that explore water chemistry including arsenic and bacteriological contamination, water salinisation, water-energy-food nexus in irrigated agriculture, and groundwater depletion resulting from irrigation, rapid urbanization, and climate change. Although the Research Topic collection does not include case studies from all the Asian Megadeltas shown in Figure 1, the relevance of multi-hazard driven water risks to all deltaic and river floodplain environments around the world is well-illustrated.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2024

A Novel Membrane-Associated Protein Aids Bacterial Colonization of Maize

The soil environment affected by plant roots and their exudates, termed the rhizosphere, significantly impacts crop health and is an attractive target for engineering desirable agricultural traits. Engineering microbes in the rhizosphere is one approach to improving crop yields that directly minimizes the number of genetic modifications made to plants. Soil microbes have the potential to assist with nutrient acquisition, heat tolerance, and drought response if they can persist in the rhizosphere in the correct numbers. Unfortunately, the mechanisms by which microbes adhere and persist on plant roots are poorly understood, limiting their application. This study examined the membrane proteome shift upon adherence to roots in two bacteria of interest, Klebsiella variicola and Pseudomonas putida. From this surface proteome data, we identified a novel membrane protein from a non-laboratory isolate of P. putida that increases binding to maize roots using unlabeled proteomics. When this protein was moved from the environmental isolate to a common lab strain (P. putida KT2440), we observed increased binding capabilities of P. putida KT2440 to both abiotic mimic surfaces and maize roots. We observed a similar increased binding capability to maize roots when the protein was heterologously expressed in K. variicola and Stutzerimonas stutzeri. With the discovery of this novel binding protein, we outline a strategy for harnessing natural selection and wild isolates to build more persistent strains of bacteria for field applications and plant growth promotion.

rhizosphere, colonization, membrane proteome, plan↗
DOE OSTI2018

Structure of the peptidoglycan polymerase RodA resolved by evolutionary coupling analysis

The shape, elongation, division and sporulation (SEDS) proteins are a large family of ubiquitous and essential transmembrane enzymes with critical roles in bacterial cell wall biology. The exact function of SEDS proteins was for a long time poorly understood, but recent work has revealed that the prototypical SEDS family member RodA is a peptidoglycan polymerase—a role previously attributed exclusively to members of the penicillin-binding protein family4. This discovery has made RodA and other SEDS proteins promising targets for the development of next-generation antibiotics. However, little is known regarding the molecular basis of SEDS activity, and no structural data are available for RodA or any homologue thereof. Here we report the crystal structure of Thermus thermophilus RodA at a resolution of 2.9 Å, determined using evolutionary covariance-based fold prediction to enable molecular replacement. The structure reveals a ten-pass transmembrane fold with large extracellular loops, one of which is partially disordered. Here, the protein contains a highly conserved cavity in the transmembrane domain, reminiscent of ligand-binding sites in transmembrane receptors. Mutagenesis experiments in Bacillus subtilis and Escherichia coli show that perturbation of this cavity abolishes RodA function both in vitro and in vivo, indicating that this cavity is catalytically essential. These results provide a framework for understanding bacterial cell wall synthesis and SEDS protein function.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

A unique class of Zn 2+ -binding serine-based PBPs underlies cephalosporin resistance and sporogenesis in Clostridioides difficile

Treatment with β-lactam antibiotics, particularly cephalosporins, is a major risk factor for Clostridioides difficile infection. These broad-spectrum antibiotics irreversibly inhibit penicillin-binding proteins (PBPs), which are serine-based enzymes that assemble the bacterial cell wall. However, C. difficile has four different PBPs (PBP1-3 and SpoVD) with various roles in growth and spore formation, and their specific links to β-lactam resistance in this pathogen are underexplored. Here, we show that PBP2 (known to be essential for vegetative growth) is the primary bactericidal target for β-lactams in C. difficile. PBP2 is insensitive to cephalosporin inhibition, and this appears to be the main basis for cephalosporin resistance in this organism. We determine crystal structures of C. difficile PBP2, alone and in complex with β-lactams, revealing unique features including ligand-induced conformational changes and an active site Zn 2+ -binding motif that influences β-lactam binding and protein stability. The Zn 2+ -binding motif is also present in C. difficile PBP3 and SpoVD (which are known to be essential for sporulation), as well as in other bacterial taxa including species living in extreme environments and the human gut. We speculate that this thiol-containing motif and its cognate Zn 2+ might function as a redox sensor to regulate cell wall synthesis for survival in adverse or anaerobic environments.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Conformational restriction shapes the inhibition of a multidrug efflux adaptor protein

Abstract Membrane efflux pumps play a major role in bacterial multidrug resistance. The tripartite multidrug efflux pump system from Escherichia coli , AcrAB-TolC, is a target for inhibition to lessen resistance development and restore antibiotic efficacy, with homologs in other ESKAPE pathogens. Here, we rationalize a mechanism of inhibition against the periplasmic adaptor protein, AcrA, using a combination of hydrogen/deuterium exchange mass spectrometry, cellular efflux assays, and molecular dynamics simulations. We define the structural dynamics of AcrA and find that an inhibitor can inflict long-range stabilisation across all four of its domains, whereas an interacting efflux substrate has minimal effect. Our results support a model where an inhibitor forms a molecular wedge within a cleft between the lipoyl and αβ barrel domains of AcrA, diminishing its conformational transmission of drug-evoked signals from AcrB to TolC. This work provides molecular insights into multidrug adaptor protein function which could be valuable for developing antimicrobial therapeutics.

59 BASIC BIOLOGICAL SCIENCES↗