Molecular mechanism of the one-component regulator RccR on bacterial metabolism and virulence.

The regulation of carbon metabolism and virulence is critical for the rapid adaptation of pathogenic bacteria to host conditions. In Pseudomonas aeruginosa, RccR is a transcriptional regulator of genes involved in primary carbon metabolism and is associated with bacterial resistance and virulence, although the exact mechanism is unclear. Our study demonstrates that PaRccR is a direct repressor of the transcriptional regulator genes mvaU and algU. Biochemical and structural analyses reveal that PaRccR can switch its DNA recognition mode through conformational changes triggered by KDPG binding or release. Mutagenesis and functional analysis underscore the significance of allosteric communication between the SIS domain and the DBD domain. Our findings suggest that, despite its overall structural similarity to other bacterial RpiR-type regulators, RccR displays a more complex regulatory element binding mode induced by ligands and a unique regulatory mechanism.

Alzheimer's disease early screening and staged detection with plasma proteome using machine learning and convolutional neural network.

Alzheimer's disease (AD) stands as the prevalent progressive neurodegenerative disease, precipitating cognitive impairment and even memory loss. Amyloid biomarkers have been extensively used in the diagnosis of AD. However, amyloid proteins offer limited information about the disease process and accurate diagnosis depends on the presence of a substantial accumulation of amyloid deposition which significantly impedes the early screening of AD. In this study, we have combined plasma proteomics with an ensemble learning model (CatBoost) to develop a cost-effective and non-invasive diagnostic method for AD. A longitudinal panel has been identified that can serve as reliable biomarkers across the entire progression of AD. Simultaneously, we have developed a neural network algorithm that utilizes plasma proteins to detect stages of Alzheimer's disease. Based on the developed longitudinal panel, the CatBoost model achieved an area under the operating curve of at least 0.90 in distinguishing mild cognitive impairment from cognitively normal. The neural network model was utilized for the detection of three stages of AD, and the results demonstrated that the neural network model exhibited an accuracy as high as 0.83, surpassing that of the traditional machine learning model.

Advances in the adenylation domain: discovery of diverse non-ribosomal peptides.

Non-ribosomal peptide synthetases are mega-enzyme assembly lines that synthesize many clinically useful compounds. As a gatekeeper, they have an adenylation (A)-domain that controls substrate specificity and plays an important role in product structural diversity. This review summarizes the natural distribution, catalytic mechanism, substrate prediction methods, and in vitro biochemical analysis of the A-domain. Taking genome mining of polyamino acid synthetases as an example, we introduce research on mining non-ribosomal peptides based on A-domains. We discuss how non-ribosomal peptide synthetases can be engineered based on the A-domain to obtain novel non-ribosomal peptides. This work provides guidance for screening non-ribosomal peptide-producing strains, offers a method to discover and identify A-domain functions, and will accelerate the engineering and genome mining of non-ribosomal peptide synthetases. KEY POINTS: • Introducing adenylation domain structure, substrate prediction, and biochemical analysis methods • Advances in mining homo polyamino acids based on adenylation domain analysis • Creating new non-ribosomal peptides by engineering adenylation domains.

Chaperone-tip adhesin complex is vital for synergistic activation of CFA/I fimbriae biogenesis.

Colonization factor CFA/I defines the major adhesive fimbriae of enterotoxigenic Escherichia coli and mediates bacterial attachment to host intestinal epithelial cells. The CFA/I fimbria consists of a tip-localized minor adhesive subunit, CfaE, and thousands of copies of the major subunit CfaB polymerized into an ordered helical rod. Biosynthesis of CFA/I fimbriae requires the assistance of the periplasmic chaperone CfaA and outer membrane usher CfaC. Although the CfaE subunit is proposed to initiate the assembly of CFA/I fimbriae, how it performs this function remains elusive. Here, we report the establishment of an in vitro assay for CFA/I fimbria assembly and show that stabilized CfaA-CfaB and CfaA-CfaE binary complexes together with CfaC are sufficient to drive fimbria formation. The presence of both CfaA-CfaE and CfaC accelerates fimbria formation, while the absence of either component leads to linearized CfaB polymers in vitro. We further report the crystal structure of the stabilized CfaA-CfaE complex, revealing features unique for biogenesis of Class 5 fimbriae.

Evaluating the efficiency of three methods to clean and disinfect screw- and cement-retained prostheses.

STATEMENT OF PROBLEM: Screw- and cement-retained prostheses (SCRPs) may be contaminated during fabrication in a dental laboratory, leading to mechanical and biological complications related to the implant treatment. Studies that explored methods to efficiently and conveniently clean and disinfect SCRPs are sparse. PURPOSE: The purpose of this clinical study was to compare the efficiency of 3 methods to remove contaminants and microorganisms present on the surface of an SCRP. MATERIAL AND METHODS: Forty-eight 1-unit SCRPs fabricated in a dental laboratory were randomly divided into 3 groups: wiping, soaking, or ultrasonic cleaning. The presence of contaminants was determined by scanning electron microscopy, and microbial cells were cultured before and after treatment. Bacterial colony-forming units (CFUs) on the surface of the SCRPs and contamination density at the implant-abutment interface and emergence profile area were assessed. Statistical tests including ANCOVA were used to compare the efficiency of different methods before and after treatment (α=.05). RESULTS: Significant differences in contamination density were noted during the treatment at the implant-abutment interface and at the emergence profile area in the 3 groups (P<.05), but no significant differences were observed in the number of CFUs (P>.05). There were significant differences among the 3 methods for cleaning efficiency both at the implant-abutment interface (P=.023) and the emergence profile area (P=.038). At the implant-abutment interface, the contamination density after treatment was lower in the ultrasonic cleaning group than that in the soaking group (P=.007), whereas at the emergence profile area, the contamination density after treatment was lower in the ultrasonic cleaning group than that in the wiping group (P=.019) and the soaking group (P=.048). CONCLUSIONS: All 3 treatment methods reduced contaminants on the SCRP surface, but ultrasonic cleaning yielded the most favorable results. However, none of the methods provided additional disinfection for SCRPs previously disinfected by ozone and UV in a dental laboratory.

Pseudomonas aeruginosa antitoxin HigA functions as a diverse regulatory factor by recognizing specific pseudopalindromic DNA motifs.

Type II toxin-antitoxin (TA) systems modulate many essential cellular processes in prokaryotic organisms. Recent studies indicate certain type II antitoxins also transcriptionally regulate other genes, besides neutralizing toxin activity. Herein, we investigated the diverse transcriptional repression properties of type II TA antitoxin PaHigA from Pseudomonas aeruginosa. Biochemical and functional analyses showed that PaHigA recognized variable pseudopalindromic DNA sequences and repressed expression of multiple genes. Furthermore, we presented high resolution structures of apo-PaHigA, PaHigA-PhigBA and PaHigA-Ppa2440 complex, describing how the rearrangements of the HTH domain accounted for the different DNA-binding patterns among HigA homologues. Moreover, we demonstrated that the N-terminal loop motion of PaHigA was associated with its apo and DNA-bound states, reflecting a switch mechanism regulating HigA antitoxin function. Collectively, this work extends our understanding of how the PaHigB/HigA system regulates multiple metabolic pathways to balance the growth and stress response in P. aeruginosa and could guide further development of anti-TA oriented strategies for pathogen treatment.

Fully Solid-State Graphene Transistors with Striking Homogeneity and Sensitivity for the Practicalization of Single-Device Electronic Bioassays.

To break through a critical barrier in the practical application of graphene biosensors, namely, device-to-device performance inhomogeneity, this work presents a novel scenario employing a fully solid-state (FSS) transistor configuration. Herein, the graphene sensing unit is completely encapsulated by a high-κ solid dielectric material, which isolates the sensing unit from solution contaminants and thus homogeneously maintains the extraordinary carrier mobility of pristine graphene in batch-made devices. To create an interface sensitive to biomolecular interactions based on the FSS configuration, a metallic floating gate functionalized by conductive mercapto-phenyl molecular linkers is defined on the top-layer solid dielectric. As the solid dielectric layer beneath the metal floating gate enables a higher capacitive gating efficiency than the regular graphene-solution electrical double layer (EDL) interface, the overall transistor amplification gain is further enhanced. As a proof of principle, a label-free DNAzymatic bioassay of Pb2+ is conducted. Without the traditional one-by-one device normalization, an excellent concentration detection limit of 929.8 fM is achieved, which is almost 2 orders of magnitude lower than that in existing works. The FSS configuration allows enhanced sensitivity and homogeneity, thereby providing new developmental guidelines for graphene biosensors beyond the laboratory investigation stage. Additionally, it has the potential to be universally applicable for cost-efficient single-device bioassays.

Classification of Fatigue Phases in Healthy and Diabetic Adults Using Wearable Sensor.

Fatigue is defined as "a loss of force-generating capacity" in a muscle that can intensify tremor. Tremor quantification can facilitate early detection of fatigue onset so that preventative or corrective controls can be taken to minimize work-related injuries and improve the performance of tasks that require high-levels of accuracy. We focused on developing a system that recognizes and classifies voluntary effort and detects phases of fatigue. The experiment was designed to extract and evaluate hand-tremor data during the performance of both rest and effort tasks. The data were collected from the wrist and finger of the participant's dominant hand. To investigate tremor, time, frequency domain features were extracted from the accelerometer signal for segments of 45 and 90 samples/window. Analysis using advanced signal processing and machine-learning techniques such as decision tree, k-nearest neighbor, support vector machine, and ensemble classifiers were applied to discover models to classify rest and effort tasks and the phases of fatigue. Evaluation of the classifier's performance was assessed based on various metrics using 5-fold cross-validation. The recognition of rest and effort tasks using an ensemble classifier based on the random subspace and window length of 45 samples was deemed to be the most accurate (96.1%). The highest accuracy (~98%) that distinguished between early and late fatigue phases was achieved using the same classifier and window length.

Methodological Approaches and Recommendations for Functional Near-Infrared Spectroscopy Applications in HF/E Research.

OBJECTIVE: The objective of this study was to systematically document current methods and protocols employed when using functional near-infrared spectroscopy (fNIRS) techniques in human factors and ergonomics (HF/E) research and generate recommendations for conducting and reporting fNIRS findings in HF/E applications. METHOD: A total of 1,687 articles were identified through Ovid-MEDLINE, PubMed, Web of Science, and Scopus databases, of which 37 articles were included in the review based on review inclusion/exclusion criteria. RESULTS: A majority of the HF/E fNIRS investigations were found in transportation, both ground and aviation, and in assessing cognitive (e.g., workload, working memory) over physical constructs. There were large variations pertaining to data cleaning, processing, and analysis approaches across the studies that warrant standardization of methodological approaches. The review identified major challenges in transparency and reporting of important fNIRS data collection and analyses specifications that diminishes study replicability, introduces potential biases, and increases likelihood of inaccurate results. As such, results reported in existing fNIRS studies need to be cautiously approached. CONCLUSION: To improve the quality of fNIRS investigations and/or to facilitate its adoption and integration in different HF/E applications, such as occupational ergonomics and rehabilitation, recommendations for fNIRS data collection, processing, analysis, and reporting are provided.

Structural and Functional Insights into PpgL, a Metal-Independent ß-Propeller Gluconolactonase That Contributes to Pseudomonas aeruginosa Virulence.

Biofilm formation is a critical determinant in the pathopoiesis of Pseudomonas aeruginosa It could significantly increase bacterial resistance to drugs and host defense. Thus, inhibition of biofilm matrix production could be regarded as a promising attempt to prevent colonization of P. aeruginosa and the subsequent infection. PpgL, a periplasmic gluconolactonase, has been reported to be involved in P. aeruginosa quorum-sensing (QS) system regulation. However, the detailed function and catalysis mechanism remain elusive. Here, the crystal structure of PpgL is described in the current study, along with biochemical analysis, revealing that PpgL is a typical ß-propeller enzyme with unique metal-independent lactone hydrolysis activity. Consequently, comparative analysis of seven-bladed propeller lactone-catalyzing enzymes and mutagenesis studies identify the critical sites which contribute to the diverse catalytic and substrate recognition functions. In addition, the reduced biofilm formation and attenuated invasion phenotype resulting from deletion of ppgL confirm the importance of PpgL in P. aeruginosa pathogenesis. These results suggest that PpgL is a potential target for developing new agents against the diseases caused by P. aeruginosa.

Mechanistic insights into the allosteric regulation of Pseudomonas aeruginosa aspartate kinase.

In plants and microorganisms, aspartate kinase (AK) catalyzes an initial commitment step of the aspartate family amino acid biosynthesis. Owing to various structural organizations, AKs from different species show tremendous diversity and complex allosteric controls. We report the crystal structure of AK from Pseudomonas aeruginosa (PaAK), a typical α2ß2 hetero-tetrameric enzyme, in complex with inhibitory effectors. Distinctive features of PaAK are revealed by structural and biochemical analyses. Essentially, the open conformation of Lys-/Thr-bound PaAK structure clarifies the inhibitory mechanism of α2ß2-type AK. Moreover, the various inhibitory effectors of PaAK have been identified and a general amino acid effector motif of AK family is described.

Monolayer Molybdenum Disulfide Transistors with Single-Atom-Thick Gates.

Two-dimensional transition-metal dichalcogenides (TMDs) are unique candidates for the development of next-generation electronic devices. However, the large contact resistance between metal and the monolayer TMDs have significantly limited the devices' performance. Also, the integration of ultrathin high- k dielectric layers with TMDs remains difficult due to the lack of dangling bonds on the surface of TMDs. We present monolayer molybdenum disulfide field-effect transistors with bottom local gates consisting of monolayer graphene. The atomic-level thickness and surface roughness of graphene facilitate the growth of high-quality ultrathin HfO2 and suppress gate leakage. Strong displacement fields above 8 V/nm can be applied using a single graphene gate to electrostatically dope the MoS2, which reduces the contact resistances between Ni and monolayer MoS2 to 2.3 kΩ·µm at low gate voltages. The devices exhibit excellent switching characteristics including a near-ideal subthreshold slope of 64 millivolts per decade, low threshold voltage (∼0.5 V), high channel conductance (>100 µS/µm), and low hysteresis. Scaled devices with 50 and 14 nm channels as well as ultrathin (5 nm) gate dielectrics show effective immunity to short-channel effects. The device fabricated on flexible polymeric substrate also exhibits high performance and has a fully transparent channel region that is desirable in optical-related studies and practical applications.

A graphene aptasensor for biomarker detection in human serum.

This paper presents an affinity graphene nanosensor for detection of biomarkers in undiluted and non-desalted human serum. The affinity nanosensor is a field-effect transistor in which graphene serves as the conducting channel. The graphene surface is sequentially functionalized with a nanolayer of the polymer polyethylene glycol (PEG) and a biomarker-specific aptamer. The aptamer is able to specifically bind with and capture unlabeled biomarkers in serum. A captured biomarker induces a change in the electric conductivity of the graphene, which is measured in a buffer of optimally chosen ionic strength to determine the biomarker concentration. The PEG layer effectively rejects nonspecific adsorption of background molecules in serum while still allowing the aptamer to be readily accessible to serum-borne biomarkers and increases the effective Debye screening length on the graphene surface. Thus, the aptamer-biomarker binding sensitively changes the graphene conductivity, thereby achieving specific and label-free detection of biomarkers with high sensitivity and without the need to dilute or desalt the serum. Experimental results demonstrate that the graphene nanosensor is capable of specifically capturing human immunoglobulin E (IgE), used as a representative biomarker, in human serum in the concentration range of 50 pM-250 nM, with a resolution of 14.5 pM and a limit of detection of 47 pM.

Enantioselective Biosynthesis of l-Phenyllactic Acid by Whole Cells of Recombinant Escherichia coli.

BACKGROUND: l-Phenyllactic acid (l-PLA)-a valuable building block in the pharmaceutical and chemical industry-has recently emerged as an important monomer in the composition of the novel degradable biocompatible material of polyphenyllactic acid. However, both normally chemically synthesized and naturally occurring phenyllactic acid are racemic, and the product yields of reported l-PLA synthesis processes remain unsatisfactory. METHODS: We developed a novel recombinant Escherichia coli strain, co-expressing l-lactate dehydrogenase (l-LDH) from Lactobacillus plantarum subsp. plantarum and glucose dehydrogenase (GDH) from Bacillus megaterium, to construct a recombinant oxidation/reduction cycle for whole-cell biotransformation of phenylpyruvic acid (PPA) into chiral l-PLA in an enantioselective and continuous manner. RESULTS: During fed-batch bioconversion with intermittent PPA feeding, l-PLA yield reached 103.8 mM, with an excellent enantiomeric excess of 99.7%. The productivity of l-PLA was as high as 5.2 mM·h-1 per OD600 (optical density at 600 nm) of whole cells. These results demonstrate the efficient production of l-PLA by the one-pot biotransformation system. Therefore, this stereoselective biocatalytic process might be a promising alternative for l-PLA production.

Dietary intake of manganese and the risk of the metabolic syndrome in a Chinese population.

Animal studies have suggested that Mn might be associated with some components of the metabolic syndrome (MetS). A few epidemiological studies have assessed dietary Mn intake and its association with the risk of the MetS and its components among Chinese adults. In this study, we assessed daily dietary Mn intake and its relationship with MetS risk among Chinese adults in Zhejiang Province using data from the 5th Chinese National Nutrition and Health Survey (2010-2012). A total of 2111 adults were included. Dietary Mn intake was assessed using 3-d 24-h dietary recalls; health-related data were obtained by questionnaire surveys, physical examinations and laboratory assessments. The mean intake of Mn was 6·07 (sd 2·94) mg/d for men (n 998) and 5·13 (sd 2·65) mg/d for women (n 1113). Rice (>42 %) was the main food source of Mn. The prevalence of the MetS was 28·0 % (590/2111). Higher Mn intake was associated with a decreased risk of the MetS in men (Q4 v. Q1 OR 0·62; 95 % CI 0·42, 0·92; P trend=0·043) but an increased risk in women (Q4 v. Q1 OR 1·56; 95 % CI 1·02, 2·45; P trend=0·078). In addition, Mn intake was inversely associated with abdominal obesity (P trend=0·016) and hypertriacylglycerolaemia (P trend=0·029) in men, but positively associated with low HDL-cholesterol in both men (P trend=0·003) and women (P trend<0·001). Our results suggest that higher Mn intakes may be protective against the MetS in men. The inverse association between Mn intake and the MetS in women might be due to the increased risk for low HDL-cholesterol.

Endogenic oxidative stress response contributes to glutathione over-accumulation in mutant Saccharomyces cerevisiae Y518.

Mechanisms of glutathione (GSH) over-accumulation in mutant Saccharomyces cerevisiae Y518 screened by ultraviolet and nitrosoguanidine-induced random mutagenesis were studied. Y518 accumulated higher levels of GSH and L-cysteine than its wild-type strain. RNA-Seq and pathway enrichment analysis indicated a difference in the expression of key genes involved in cysteine production, the GSH biosynthesis pathway, and antioxidation processes. GSH1, MET17, CYS4, GPX2, CTT1, TRX2, and SOD1 and the transcriptional activators SKN7 and YAP1 were up-regulated in the mutant. Moreover, Y518 showed a dysfunctional respiratory chain resulting from dramatically weakened activity of complex III and significant elevation of intracellular reactive oxygen species (ROS) levels. The supplementation of antimycin A in the culture of the parent strain showed equivalent changes of ROS and GSH level. This study indicates that defective complex III prompts abundant endogenic ROS generation, which triggers an oxidative stress response and upregulation of gene expression associated with GSH biosynthesis. This finding may be helpful for developing new strategies for GSH fermentation process optimization or metabolic engineering.

Enhancement of phenyllactic acid biosynthesis by recognition site replacement of D-lactate dehydrogenase from Lactobacillus pentosus.

OBJECTIVES: The Tyr52 residue of D-lactate dehydrogenase (D-LDH) from Lactobacillus pentosus was replaced with small hydrophobic residues and overexpressed in E. coli BL21 (DE3) to enhance 3-phenyllactic acid (PLA) synthesis by whole-cell catalyst. RESULTS: Escherichia coli pET-28a-d-ldh produced 12.2 g PLA l(-1) in 3 h, with a molar conversion rate of 61 %, while E. coli pET-28a-d-ldh (Y52V) produced 15.6 g PLA l(-1), with a molar conversion rate of 77 %. This study demonstrates the feasibility of using engineered E. coli for PLA production from phenylpyruvate (PPA) and showed that site-directed mutagenesis of d-ldh markedly improved PLA yield and substrate conversion rate. CONCLUSION: This biocatalytic system is a promising platform for PLA production from PPA.

Optimal Selection of Sampling Points within Sewer Networks for Wastewater-Based Epidemiology Applications.

Wastewater-based epidemiology (WBE) has great potential to monitor community public health, especially during pandemics. However, it faces substantial hurdles in pathogen surveillance through WBE, encompassing data representativeness, spatiotemporal variability, population estimates, pathogen decay, and environmental factors. This paper aims to enhance the reliability of WBE data, especially for early outbreak detection and improved sampling strategies within sewer networks. The tool implemented in this paper combines a monitoring model and an optimization model to facilitate the optimal selection of sampling points within sewer networks. The monitoring model utilizes parameters such as feces density and average water consumption to define the detectability of the virus that needs to be monitored. This allows for standardization and simplicity in the process of moving from the analysis of wastewater samples to the identification of infection in the source area. The entropy-based model can select optimal sampling points in a sewer network to obtain the most specific information at a minimum cost. The practicality of our tool is validated using data from Hildesheim, Germany, employing SARS-CoV-2 as a pilot pathogen. It is important to note that the tool's versatility empowers its extension to monitor other pathogens in the future.

Bedside electrical impedance tomography to assist the management of pulmonary embolism: A case report.

Background: Pulmonary embolism (PE) is a common worldwide disease with high mortality. Timely diagnosis and management of PE could significantly improve clinical outcomes. Electrical impedance tomography (EIT) is a novel noninvasive technique to monitor lung perfusion and help detect PE at the bedside. Here we present a case of clinical management of subsegmental PE with the help of the bilateral ventilation and perfusion(V/Q) asymmetry EIT image. Case presentation: A 72-year-old cancer patient with respiratory failure and acute kidney injury in the intensive care unit was suspected of PE based on his clinical manifestation. The contraindication of computed tomography pulmonary angiography (CTPA) for PE diagnosis prevented escalating anticoagulation therapy. Besides EIT ventilation and perfusion monitoring showed an abnormal asymmetry V/Q match between the bilateral lungs which promoted our decision to start systemic continuous anticoagulation therapy and improved the patient clinically. The following CTPA which clarified the diagnosis of PE suggests that the patient has benefited from our decision. Conclusion: For critically ill patients with suspected PE, the asymmetry of the EIT V/Q image may provide crucial objective information for clinical management.

Potential molecular mechanisms of Huangqin Tang for liver cancer treatment by network pharmacology and molecular dynamics simulations.

OBJECTIVE: This study aims to investigate the mechanism of Huangqin Tang in treating liver cancer. METHODS: Active ingredients and corresponding targets of Huangqin Tang were obtained from the Traditional Chinese Medicine Systems Pharmacology Database. Differentially expressed genes in liver cancer were identified from mRNA expression data. A protein-protein interaction (PPI) network was constructed using differentially expressed genes and Huangqin Tang targets. Random walk with restart (RWR) analysis was performed on the PPI network. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were conducted. A drug-active ingredient-gene interaction network was established, and molecular docking and molecular dynamics simulations were performed. Finally, the stability of binding between CDK1 and oroxylin was tested according to cellular thermal shift assay (CETSA). RESULTS: 160 active ingredients, 239 targets, and 1093 differentially expressed genes were identified. RWR analysis identified 10 potential targets for liver cancer. Enrichment analysis revealed protein kinase regulator activity and Steroid hormone biosynthesis as significant pathways. Molecular docking suggested a stable complex between oroxylin A and CDK1. CETSA demonstrated that the combination of oroxylin A and CDK1 increased the stability of CDK1, and the combination efficiency was high. CONCLUSION: Huangqin Tang may treat liver cancer by targeting CDK1 with oroxylin A. Protein kinase regulator activity and Steroid hormone biosynthesis pathways may play a role in liver cancer treatment with Huangqin Tang. This study provides insight into the mechanistic basis of Huangqin Tang for liver cancer treatment.