Role of N-acetylkynurenine in mediating the effect of gut microbiota on urinary tract infection: a Mendelian randomization study.

Introduction: This study explored the causal connections between gut microbiota (GM), urinary tract infection (UTI), and potential metabolite mediators using Mendelian randomization (MR). Methods: We utilized summary statistics from the most comprehensive and extensive genome-wide association studies (GWAS) available to date, including 196 bacterial traits for GM, 1,091 blood metabolites, 309 metabolite ratios, alongside UTI data from ukb-b-8814 and ebi-a-GCST90013890. Bidirectional MR analyses were conducted to investigate the causal links between GM and UTI. Subsequently, two MR analyses were performed to identify the potential mediating metabolites, followed by a two-step MR analysis to quantify the mediation proportion. Results: Our findings revealed that out of the total 15 bacterial traits, significant associations with UTI risk were observed across both datasets. Particularly, taxon g_Ruminococcaceae UCG010 displayed a causal link with a diminished UTI risk in both datasets (ukb-b-8814: odds ratio [OR] = 0.9964, 95% confidence interval [CI] = 0.9930-0.9997, P = 0.036; GCST90013890: OR = 0.8252, 95% CI = 0.7217-0.9436, P = 0.005). However, no substantial changes in g_Ruminococcaceae UCG010 due to UTI were noted (ukb-b-8814: ß = 0.51, P = 0.87; ebi-a-GCST90013890: ß = -0.02, P = 0.77). Additionally, variations in 56 specific metabolites were induced by g_Ruminococcaceae UCG010, with N-acetylkynurenine (NAK) exhibiting a causal correlation with UTI. A negative association was found between g_Ruminococcaceae UCG010 and NAK (OR: 0.8128, 95% CI: 0.6647-0.9941, P = 0.044), while NAK was positively associated with UTI risk (OR: 1.0009; 95% CI: 1.0002-1.0016; P = 0.0173). Mediation analysis revealed that the association between g_Ruminococcaceae UCG010 and UTI was mediated by NAK with a mediation proportion of 5.07%. Discussion: This MR study provides compelling evidence supporting the existence of causal relationships between specific GM taxa and UTI, along with potential mediating metabolites.

Self-cascade deoxynivalenol detoxification by an artificial enzyme with bifunctions of dehydrogenase and aldo/keto reductase from genome mining.

Due to the severe health risks for human and animal caused by the intake of toxic deoxynivalenol (DON) derived from Fusarium species, elimination DON in food and feed has been initiated as a critical issue. Enzymatic cascade catalysis by dehydrogenase and aldo-keto reductase represents a fascinating strategy for DON detoxification. Here, one quinone-dpendent alcohol dehydrogenase DADH oxidized DON into less-toxic 3-keto-DON and NADPH-dependent aldo-keto reductase AKR13B3 reduced 3-keto-DON into relatively non-toxic 3-epi-DON were identified from Devosia strain A6-243, indicating that degradation of DON on C3 are two-step sequential cascade processes. To establish the bifunctions, fusion enzyme linking DADH and AKR13B3 was successfully assembled to promote one-step DON degradations with accelerated specific activity and efficiency, resulting 93.29 % of DON removal rate in wheat sample. Three-dimensional simulation analysis revealed that the bifunctional enzyme forms an artificial intramolecular channel to minimize the distance of intermediate from DADH to AKR13B3 for two-step enzymatic reactions, and thereby accelerates this enzymatic process. As the first report of directing single step DON detoxification by an interesting bifunctional artificial enzyme, this work revealed a facile and eco-friendly approach to detoxify DON with application potential and gave valuable insights into execute other mycotoxin detoxification for ensuring food safety.

Cis-Element Engineering Promotes the Expression of Bacillus subtilis Type I L-Asparaginase and Its Application in Food.

Type I L-asparaginase from Bacillus licheniformis Z-1 (BlAase) was efficiently produced and secreted in Bacillus subtilis RIK 1285, but its low yield made it unsuitable for industrial use. Thus, a combined method was used in this study to boost BlAase synthesis in B. subtilis. First, fifteen single strong promoters were chosen to replace the original promoter P43, with PyvyD achieving the greatest BlAase activity (436.28 U/mL). Second, dual-promoter systems were built using four promoters (PyvyD, P43, PaprE, and PspoVG) with relatively high BlAase expression levels to boost BlAase output, with the engine of promoter PaprE-PyvyD reaching 502.11 U/mL. The activity of BlAase was also increased (568.59 U/mL) by modifying key portions of the PaprE-PyvyD promoter. Third, when the ribosome binding site (RBS) sequence of promoter PyvyD was replaced, BlAase activity reached 790.1 U/mL, which was 2.27 times greater than the original promoter P43 strain. After 36 h of cultivation, the BlAase expression level in a 10 L fermenter reached 2163.09 U/mL, which was 6.2 times greater than the initial strain using promoter P43. Moreover, the application potential of BlAase on acrylamide migration in potato chips was evaluated. Results showed that 89.50% of acrylamide in fried potato chips could be removed when combined with blanching and BlAase treatment. These findings revealed that combining transcription and translation techniques are effective strategies to boost recombinant protein output, and BlAase can be a great candidate for controlling acrylamide in food processing.

Structure-Function Analysis of a Quinone-Dependent Dehydrogenase Capable of Deoxynivalenol Detoxification.

The pyrroloquinoline quinone (PQQ)-dependent dehydrogenase DepA detoxifies deoxynivalenol (DON) by converting the C3-OH into a keto group. Herein, two crystal structures of DepA and its complex with PQQ were determined, together with biochemical evidence confirming the interactions of DepA with PQQ and DON and revealing a unique tyrosine residue important for substrate selection. Furthermore, four loops over the active site essential for DepA activity were identified, of which three loops were stabilized by PQQ, and the fourth loop invisible in both structures was considered important for binding DON, together constituting a lid for the active site. Preliminary engineering of the loop showed its potential for enzyme improvement. This study provides structural insights into how a PQQ-dependent dehydrogenase is equipped with the function of DON conversion and for the first time shows the necessity of a lid structure for PQQ-dependent dehydrogenase activity, laying foundation for structure-based design to enhance catalysis efficiency.

Microwave-Assisted Extraction of Multiple Trace Levels of Intermediate Metabolites for Camptothecin Biosynthesis in Camptotheca acuminata and Their Simultaneous Determination by HPLC-LTQ-Orbitrap-MS/MS and HPLC-TSQ-MS.

Camptothecin (CPT) has strong antitumor activity and is used as an anticancer therapeutic agent. To better understand and decipher the pathway of CPT biosynthesis in Camptotheca acuminata, the main purpose here was focused on creating an effective extraction strategy for a rich intermediate metabolite profile. In the present study, a 70% aqueous acetonitrile was verified as an optimal extraction solvent for microwave-assisted extraction (MAE) of metabolites by spiking experiments. Based on multi-objective optimization, the best extraction conditions of a solid-liquid ratio of 1:20, microwave power of 230 W, and a time of 4 min were achieved using a full factorial 34 experimental design. Crude extracts obtained from the shoot apex of C. acuminata using MAE have been qualitatively profiled by high-performance liquid chromatography coupled with linear ion trap quadrupole-orbitrap mass spectrometry (HPLC-LTQ-Orbitrap-MS/MS) and a HPLC triple quadrupole-MS (HPLC-TSQ-MS) analysis was conducted for their metabolite content in different tissues. CPT, and ten related metabolites and their isomers, including tryptamine, loganic acid, secologanic acid, strictosidinic acid, strictosamide, strictosamide epoxide, strictosamide diol, strictosamide ketolactam, pumiloside, and deoxypumiloside, were detected and tentatively identified. Scanning electron microscopy (SEM) imaging of the shoot apex demonstrated that severe cell disruption was evident after intensified extraction processes. The study showed the difference of metabolite profiles and the enhancement of metabolite content after microwave-pretreated techniques, and the established MAE procedure is an effective methodology to preserve valuable metabolite compounds for analysis.