Enzyme Function Research Articles

An ancient role for CYP73 monooxygenases in phenylpropanoid biosynthesis and embryophyte development.

The phenylpropanoid pathway is one of the plant metabolic pathways most prominently linked to the transition to terrestrial life, but its evolution and early functions remain elusive. Here, we show that activity of the t-cinnamic acid 4-hydroxylase (C4H), the first plant-specific step in the pathway, emerged concomitantly with the CYP73 gene family in a common ancestor of embryophytes. Through structural studies, we identify conserved CYP73 residues, including a crucial arginine, that have supported C4H activity since the early stages of its evolution. We further demonstrate that impairing C4H function via CYP73 gene inactivation or inhibitor treatment in three bryophyte species-the moss Physcomitrium patens, the liverwort Marchantia polymorpha and the hornwort Anthoceros agrestis-consistently resulted in a shortage of phenylpropanoids and abnormal plant development. The latter could be rescued in the moss by exogenous supply of p-coumaric acid, the product of C4H. Our findings establish the emergence of the CYP73 gene family as a foundational event in the development of the plant phenylpropanoid pathway, and underscore the deep-rooted function of the C4H enzyme in embryophyte biology.

Exploration of Hydrazide-Based HDAC8 PROTACs for the Treatment of Hematological Malignancies and Solid Tumors.

HDAC8 can mediate signals by using its enzymatic or nonenzymatic functions, which are expected to be critical for various types of cancer. Herein, we employed proteolysis targeting chimera (PROTAC) technology to target the enzymatic as well as the nonenzymatic functions of HDAC8. A potent and selective HDAC8 PROTAC Z16 (CZH-726) with low nanomolar DC50 values in various cell lines was identified. Interestingly, Z16 induced structural maintenance of chromosomes protein 3 (SMC3) hyperacetylation at low concentrations and histone hyperacetylation at high concentrations, which can be explained by HDAC8 degradation and off-target HDAC inhibition, respectively. Notably, Z16 potently inhibited proliferation of various cancer cell lines and the antiproliferative mechanisms proved to be cell-type-dependent, which, to a large extent, is due to off-target HDAC inhibition. In conclusion, we report a hydrazide-based HDAC8 PROTAC Z16, which can be used as a probe to investigate the biological functions of HDAC8.

Conformation-Dependent Hydrogen-Bonding Interactions in a Switchable Artificial Metalloprotein.

Hydrogen-bonding (H-bonding) interactions in metalloprotein active sites can critically regulate enzyme function. Changes in the protein structure triggered by interplay with substrates, products, and partner proteins are often translated to the metallocofactor by way of specific changes in H-bond networks connected to the active site. However, the complexities of metalloprotein architecture and mechanism often preclude our ability to define the precise molecular interactions giving rise to these intricate regulatory pathways. To address this shortcoming, we have developed conformationally switchable artificial metalloproteins (swArMs) in which allosteric Gln-binding triggers protein conformational changes that impact the microenvironment surrounding an installed metallocofactor. Herein, we report a combined structural, spectroscopic, and computational approach to enhance the conformation-dependent changes in H-bond interactions surrounding the metallocofactor site of a swArM. Structure-informed molecular dynamics simulations were employed to predict point mutations that could enhance active site H-bond interactions preferentially in the Gln-bound holo-conformation of the swArM. Testing our predictions via the unique infrared spectral signals associated with the metallocofactor site, we have identified three key residues capable of imparting conformational control over the metallocofactor microenvironment. The resultant swArMs not only model biologically relevant structural regulation but also provide an enhanced Gln-responsive biological probe to be leveraged in future biosensing applications.

Long-term application of organic matter improves soil properties and plant growth-promoting bacteria in soil communities of oil palm plantation

ABSTRACT Oil palm (Elaeis guineensis) is a major contributor to global vegetable oil production; however, ensuring its sustainability remains a critical challenge, particularly concerning soil health. In this study, we investigated the impact of long-term organic matter application as part of good soil management (GSM) practices on oil palm plantations and compared it with poor soil management practices to determine the presence of plant growth-promoting bacteria (PGPB) in soil communities. The ten-years regular application of organic matter to the soil in the GSM plots led to notable improvements in soil chemical properties, including total organic carbon, total nitrogen, available phosphorus, available potassium, and cation exchange capacity. Metagenomic analysis revealed a significantly higher abundance of beneficial microbial species exclusively found in GSM plots, supporting oil palm growth. Furthermore, a novel finding emerged from this study, as it successfully predicted the metabolic function of PGPB in soil communities using PICRUST2 provided by the soil microbiome. PICRUST2 analysis indicated that the long-term application of organic matter in GSM plots increased functional enzymes related to PGP activities, such as nitrogen fixation, phosphate solubilization, potassium solubilization, and phytohormone synthesis. This study underscores the significance of implementing GSM practices in oil palm plantations by incorporating eco-friendly materials, such as organic matter, to enhance soil health and fertility and ensure oil palm sustainability.

The synergistic effects of dietary protein and stocking density on performance, digestive enzymes, and hemato-biochemistry parameters of juvenile striped catfish (Pangasianodon hypophthalmus)

This experiment was designed to examine the impacts of different dietary protein levels (DPLs) and their interaction on the growth and some physiological indices of juvenile striped catfish, Pangasianodon hypophthalmus, reared at different stocking densities (SDs) for 15 weeks. A total of 1440 striped catfish (initial weight = 10.0 ± 2.2 g) were randomly allocated into six equal treated groups (three replicates per treated group). Fish were fed two DPLs of 25 % and 30 % and reared under different SDs of 20, 40, and 60 fish/m3. The obtained results indicated that with increasing DPL, growth parameters, and digestive enzyme activities, the liver function enzyme activities, serum protein constituents' levels, and concentrations of thyroid hormones significantly increased, while levels of serum glucose, total cholesterol, and triglycerides remarkably decreased. On the other hand, increasing SD negatively affects growth, digestive and liver enzyme activities, some hematological parameters, and fish-body chemical analysis. The interaction between DPLs and different SDs significantly improved the most estimated parameters with increasing DPLs despite intensifying SDs. In general, afforded striped catfish diets containing 30 % DPL and stocked at 20 fish/m3 presented the best performance, consumed feed efficiency, and health status. Thus, the interaction between DPLs and SDs demonstrated the favorable benefits of increasing DPLs by diminishing some of the adverse impacts generated by high SDs, which may subsequently lead to high production and increased revenues for fish farmers.

Missense variants in CMS22 patients reveal that PREPL has both enzymatic and non-enzymatic functions.

Congenital myasthenic syndrome-22 (CMS22, OMIM 616224) is a rare genetic disorder caused by deleterious genetic variation in the prolyl endopeptidase-like (PREPL) gene. Previous reports have described patients with deletions and nonsense variants in PREPL, but nothing is known about the effect of missense variants in the pathology of CMS22. In this study, we have functionally characterized missense variants in PREPL from three CMS22 patients, all with hallmark phenotypes. Biochemical evaluation revealed that these missense variants do not impair hydrolase activity, thereby challenging the conventional diagnostic criteria and disease mechanism. Structural analysis showed that the variants affect regions most likely involved in intra-protein or protein-protein interactions. Indeed, binding to a selected group of known interactors was differentially reduced for the three mutants. The importance of non-hydrolytic functions of PREPL was investigated in catalytically inactive PREPL p.Ser559Ala cell lines which showed that hydrolytic activity of PREPL is needed for normal mitochondrial function but not for regulating AP1-mediated transport in the trans-Golgi network. In conclusion, these studies showed that CMS22 can be caused not only by deletion and truncation of PREPL but also by missense variants that do not necessarily result in a loss of hydrolytic activity of PREPL.

Are carriers unaffected? A literature review of metabolic and health outcomes among genetic carriers of phenylketonuria (PKU).

Phenylketonuria (PKU) is an autosomal recessive genetic condition that results in reduced enzymatic functioning within the phenylalanine hydroxylase (PAH) pathway, which is involved in the metabolism of Phenylalanine (Phe) into Tyrosine (Tyr). Without dietary intervention, individuals with PKU exhibit significantly elevated levels of Phe, which is presumed to cause severe neurological dysfunction and other associated health risks. Carriers of PKU are heterozygotes for a PAH gene mutation and are typically described in the literature as "unaffected." However, decades of existing research challenges this classical thinking and it is plausible that these individuals currently classified as carriers may present with an intermediate phenotype or may be "moderately affected." The purpose of this scoping review was to explore this hypothesis further, by searching for and summarizing existing literature on metabolism and health outcomes among PKU carriers. Preliminary research has suggested that some PKU carriers exhibit reduced PAH enzyme function, and related, elevated circulating Phe levels compared to non-carriers. In addition, Phe dosing trials have further demonstrated that carriers have increased Phe levels and decreased Tyr levels compared to non-carriers. Because of these metabolic perturbations, it is biologically plausible for carriers to experience an intermediate phenotype in terms of metabolic consequences and clinical outcomes. While these outcomes have yet to be thoroughly explored, early research has found associations between PKU carrier status and lower IQs as well as decreased executive functioning, memory, processing speed, and inhibitory control. The PAH pathway is also involved in melanogenesis and research has demonstrated increased melanoma risk among PKU carriers. However, there are many limitations to this research and thus whether or not carriers are clinically impacted cannot yet be conclusively determined. Overall, while preliminary research suggests a possible intermediate phenotype among PKU carriers, the current available research is limited and PKU carriers are still clinically considered "unaffected." This review outlines the current literature while discussing future research endeavours related to the metabolism and health of PKU carriers.

Licorice and liver function in patients with primary liver disease: A systematic review and meta-analysis of RCTs.

Licorice (Glycyrrhiza spp.) has been a cornerstone of traditional Chinese and Japanese medicine. This systematic review and meta-analysis aimed to evaluate the efficacy of licorice formulations, alone or in combination with other herbs, on liver function enzymes in patients with primary liver disease. We systematically searched MEDLINE, Embase, Scopus, Web of Science, and Cochrane Library up to April 2024. Randomized controlled trials (RCTs) comparing the effects of Glycyrrhiza spp. preparations versus placebo or standard of care controls were included. Standard Cochrane methods were used to extract data and appraise eligible studies. A total of 15 RCTs, involving 1367 participants, were included in the analysis. The studies varied widely in geographical location, duration, and licorice preparations used. Licorice significantly reduced alanine aminotransferase (ALT) by 15.63 U/L (95% CI: -25.08, -6.18; p = 0.001) and aspartate aminotransferase (AST) by 7.37 U/L (95% CI: -13.13, -1.61; p = 0.01) compared to control groups. Subgroup analyses revealed that purified glycyrrhizic acid compounds were particularly effective, showing greater reductions in ALT and AST without significant heterogeneity. Although licorice treatment did not significantly impact gamma-glutamyl transferase and total bilirubin (TBIL) levels overall, specific licorice-herb preparations did show a notable reduction in TBIL. The safety profile of licorice was consistent with known side effects, predominantly mild and related to its mineralocorticoid effects. Despite heterogeneity and potential language bias, the findings suggest that licorice can enhance liver function. Further studies should standardize licorice preparations and explore its role in multifaceted herbal formulations to better understand its hepatoprotective mechanisms.

Machine learning-guided co-optimization of fitness and diversity facilitates combinatorial library design in enzyme engineering

The effective design of combinatorial libraries to balance fitness and diversity facilitates the engineering of useful enzyme functions, particularly those that are poorly characterized or unknown in biology. We introduce MODIFY, a machine learning (ML) algorithm that learns from natural protein sequences to infer evolutionarily plausible mutations and predict enzyme fitness. MODIFY co-optimizes predicted fitness and sequence diversity of starting libraries, prioritizing high-fitness variants while ensuring broad sequence coverage. In silico evaluation shows that MODIFY outperforms state-of-the-art unsupervised methods in zero-shot fitness prediction and enables ML-guided directed evolution with enhanced efficiency. Using MODIFY, we engineer generalist biocatalysts derived from a thermostable cytochrome c to achieve enantioselective C-B and C-Si bond formation via a new-to-nature carbene transfer mechanism, leading to biocatalysts six mutations away from previously developed enzymes while exhibiting superior or comparable activities. These results demonstrate MODIFY’s potential in solving challenging enzyme engineering problems beyond the reach of classic directed evolution.

OCT1 (SLC22A1) transporter kinetics and regulation in primary human hepatocyte 3D spheroids

3D spheroids of primary human hepatocytes (3D PHH) retain a differentiated phenotype with largely conserved metabolic function and proteomic fingerprint over weeks in culture. As a result, 3D PHH are gaining importance as a model for mechanistic liver homeostasis studies and in vitro to in vivo extrapolation (IVIVE) in drug discovery. However, the kinetics and regulation of drug transporters have not yet been assessed in 3D PHH. Here, we used organic cation transporter 1 (OCT1/SLC22A1) as a model to study both transport kinetics and the long-term regulation of transporter activity via relevant signalling pathways. The kinetics of the OCT1 transporter was studied using the fluorescent model substrate 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP+) and known OCT1 inhibitors in individual 3D PHH. For long-term studies, 3D PHH were treated with xenobiotics for seven days, after which protein expression and OCT1 function were assessed. Global proteomic analysis was used to track hepatic phenotypes as well as prototypical changes in other regulated proteins, such as P-glycoprotein and Cytochrome P450 3A4. ASP+ kinetics indicated a fully functional OCT1 transporter with a Km value of 14 ± 4.0µM as the mean from three donors. Co-incubation with known OCT1 inhibitors decreased the uptake of ASP+ in the 3D PHH spheroids by 35–52%. The long-term exposure studies showed that OCT1 is relatively stable upon activation of nuclear receptor signalling or exposure to compounds that could induce inflammation, steatosis or liver injury. Our results demonstrate that 3D PHH spheroids express physiologically relevant levels of fully active OCT1 and that its transporter kinetics can be accurately studied in the 3D PHH configuration. We also confirm that OCT1 remains stable and functional during the activation of key metabolic pathways that alter the expression and function of other drug transporters and drug-metabolizing enzymes. These results will expand the range of studies that can be performed using 3D PHH.

A zebrafish gephyrinb mutant distinguishes synaptic and enzymatic functions of Gephyrin

Gephyrin is thought to play a critical role in clustering glycine receptors at synapses within the central nervous system (CNS). The main in vivo evidence for this comes from Gephyrin (Gphn)-null mice, where glycine receptors are depleted from synaptic regions. However, these mice die at birth, possibly due to impaired molybdenum cofactor (MoCo) synthesis, an essential role Gephyrin assumes throughout an animal. This complicates the interpretation of synaptic phenotypes in Gphn-null mice and raises the question whether the synaptic and enzymatic functions of Gephyrin can be investigated separately. Here, we generated a gephyrinb zebrafish mutant, vo84, that almost entirely lacks Gephyrin staining in the spinal cord. gephyrinbvo84 mutants exhibit normal gross morphology at both larval and adult stages. In contrast to Gphn-null mice, gephyrinbvo84 mutants exhibit normal motor activity and MoCo-dependent enzyme activity. Instead, gephyrinbvo84 mutants display impaired rheotaxis and increased mortality in late development. To investigate what may mediate these defects in gephyrinbvo84 mutants, we examined the cell density of neurons and myelin in the spinal cord and found no obvious changes. Surprisingly, in gephyrinbvo84 mutants, glycine receptors are still present in the synaptic regions. However, their abundance is reduced, potentially contributing to the observed defects. These findings challenge the notion that Gephyrin is absolutely required to cluster glycine receptors at synapses and reveals a new role of Gephyrin in regulating glycine receptor abundance and rheotaxis. They also establish a powerful new model for studying the mechanisms underlying synaptic, rather than enzymatic, functions of Gephyrin.

Effects of nitrile compounds on the structure and function of soil microbial communities as revealed by metagenomes

The proliferation of nitrile mixtures has significantly exacerbated environmental pollution. This study employed metagenomic analysis to investigate the short-term effects of nitrile mixtures on soil microbial communities and their metabolic functions. It also examined the responses of indigenous microorganisms and their functional metabolic genes across various land use types to different nitrile stressors. The nitrile compound treatments in this study resulted in an increase in the abundance of Proteobacteria, Actinobacteria, and Firmicutes, while simultaneously reducing overall microbial diversity. The key genes involved in the denitrification process, namely, nirK, nosZ, and hao, were down-regulated, and NO3−-N, NO2−-N, and NH4+-N concentrations decreased by 7.7%–12.3%, 11.1%–21.3%, and 11.3%–30.9%, respectively. Notably, pond sludge samples exhibited a significant increase in the abundance of nitrogen fixation-related genes nifH, vnfK, vnfH, and vnfG following exposure to nitrile compounds. Furthermore, the fumarase gene fumD, which is responsible for catalyzing fumaric acid into malic acid in the tricarboxylic acid cycle, showed a substantial increase of 7.2–10.6-fold upon nitrile addition. Enzyme genes associated with the catechol pathway, including benB-xylY, dmpB, dmpC, dmpH, and mhpD, displayed increased abundance, whereas genes related to the benzoyl-coenzyme A pathway, such as bcrA, dch, had, oah, and gcdA, were notably reduced. In summary, complex nitrile compounds were found to significantly reduce the species diversity of soil microorganisms. Nitrile-tolerant microorganisms demonstrated the ability to degrade and adapt to nitrile pollutants by enhancing functional enzymes involved in the catechol pathway and fenugreek conversion pathway. This study offers insights into the specific responses of microorganisms to compound nitrile contamination, as well as valuable information for screening nitrile-degrading microorganisms and identifying nitrile metabolic enzymes.

GloEC: a hierarchical-aware global model for predicting enzyme function.

The annotation of enzyme function is a fundamental challenge in industrial biotechnology and pathologies. Numerous computational methods have been proposed to predict enzyme function by annotating enzyme labels with Enzyme Commission number. However, the existing methods face difficulties in modelling the hierarchical structure of enzyme label in a global view. Moreover, they haven't gone entirely to leverage the mutual interactions between different levels of enzyme label. In this paper, we formulate the hierarchy of enzyme label as a directed enzyme graph and propose a hierarchy-GCN (Graph Convolutional Network) encoder to globally model enzyme label dependency on the enzyme graph. Based on the enzyme hierarchy encoder, we develop an end-to-end hierarchical-aware global model named GloEC to predict enzyme function. GloEC learns hierarchical-aware enzyme label embeddings via the hierarchy-GCN encoder and conducts deductive fusion of label-aware enzyme features to predict enzyme labels. Meanwhile, our hierarchy-GCN encoder is designed to bidirectionally compute to investigate the enzyme label correlation information in both bottom-up and top-down manners, which has not been explored in enzyme function prediction. Comparative experiments on three benchmark datasets show that GloEC achieves better predictive performance as compared to the existing methods. The case studies also demonstrate that GloEC is capable of effectively predicting the function of isoenzyme. GloEC is available at: https://github.com/hyr0771/GloEC.

The Interplay of TLR-NFκB Signalling Pathway and Functional Immune-Related Enzymes in the Inflammatory Response of Ciona robusta

The Interplay of TLR-NFκB Signalling Pathway and Functional Immune-Related Enzymes in the Inflammatory Response of Ciona robusta

A tethering mechanism underlies Pin1-catalyzed proline cis-trans isomerization at a noncanonical site.

Proline peptide bonds naturally occur in cis conformations and isomerize to trans conformations on exchange regimes on the order of seconds to minutes. Pin1, a prolyl isomerase, catalyzes the isomerization of proline peptide bonds that contain a specific phospho-motif-a phosphorylated serine or threonine followed by a proline (pS/pT-P)-allowing for switch-like effects on target protein structure and function. One protein substrate of Pin1 is the nuclear receptor peroxisome proliferator activated receptor gamma (PPARγ), which is shown here to undergo Pin1-catalyzed isomerization at a noncanonical proline distal to a canonical pS/pT-P binding site. These studies lay the foundation for understanding the role of Pin1 in mediating PPARγ-regulated transcription and expand understanding of Pin1-catalyzed enzymatic activities and functions.

FOXM1 requires IDH1 for late genes expression in mitotic cells.

Isocitrate dehydrogenase 1 (IDH1) is a metabolic enzyme that converts isocitrate to α-ketoglutarate in cells. However, research on IDH1 is more focused on the metabolite D-2-hydroxyglutarate than the cellular roles of the IDH1 protein. Metabolic enzymes can moonlight by participating in diverse cellular processes in cancer cells. This moonlighting function of the metabolic enzymes can contribute to changes in gene expression. It is unknown whether IDH1 associates with any transcription factor. We asked whether IDH1 coordinates with forkhead box protein M1 (FOXM1) in mitotic cells to regulate late genes expression. We found that depletion of IDH1 reduces canonical FOXM1-target expression in mitotic cells. Also, IDH1 binds to FOXM1 and a subset of MuvB proteins, Lin-9 and Lin-54, in mitotic cells. Based on these observations, we suggest that IDH1 coordinates with FOXM1 in mitotic cells to regulate late genes expression.

Oxysterol binding protein (OSBP) contributes to hepatitis E virus replication

Hepatitis E virus (HEV) is a positive-sense, single-stranded RNA virus and causes primarily acute self-limiting infections. The ORF1 of the HEV genome encodes a polyprotein around 190 kDa, which contains several putative domains, including helicase and RNA-dependent RNA polymerase. The HEV-encoded helicase is a member of the superfamily 1 helicase family and possesses multiple enzymatic functions, such as RNA 5′-triphosphatase, RNA unwinding, and NTPase, which are thought to contribute to viral RNA synthesis. However, the helicase interaction with cellular proteins remains less known. Oxysterol binding protein (OSBP) is a lipid regulator that shuffles between the Golgi apparatus and the endoplasmic reticulum for cholesterol and phosphatidylinositol-4-phosphate exchange and controls the efflux of cholesterol from cells. In this study, the RNAi-mediated silencing of OSBP significantly reduced HEV replication. Further studies indicate that the HEV helicase interacted with OSBP, shown by co-immunoprecipitation and co-localization in co-transfected cells. The presence of helicase blocked OSBP preferential translocation to the Golgi apparatus. These results demonstrate that OSBP contributes to HEV replication and enrich our understanding of the HEV-cell interactions.

Structure-Guided Identification of Novel Aromatase Inhibitors Targeting Breast Carcinoma.

Aromatase inhibitors play a critical therapeutic role in treating ER+ breast cancer, especially in postmenopausal women. However, their efficacy is often limited by resistance and severe side effects. Identifying new compounds that can disrupt aromatase enzyme function is essential. In this study, structural anomalies in the aromatase enzyme were corrected through energy minimization, and the structure was validated via Ramachandran plot. We screened 170,269 natural compounds from the ASINEX Biodesign library using high-throughput screening algorithms to target the aromatase enzyme. Molecular docking identified three compounds: BDD30170158, BDE33872639, and BDE30177677, all showing stable binding interactions with the aromatase enzyme. Molecular dynamics simulations over 100 ns confirmed the conformational stability of these compounds. Although all three compounds exhibited the desired pharmacokinetic and drug metabolism properties, only one compound (BDE33872639) was identified as a non-blocker, demonstrating a reduced risk of adverse cardiac effects. This compound exhibits significant potential as a novel aromatase inhibitor, warranting further experimental research to develop it as a therapeutic option for ER+ breast cancer.

Variant analysis and PGT-M of OTC gene in a Chinese family with ornithine carbamoyltransferase deficiency

BackgroundOrnithine carbamoyltransferase deficiency (OTCD) is a kind of X-linked metabolic disease caused by a deficiency in ornithine transcarbamylase leading to urea cycle disorders. The main reason is that the OTC gene variants lead to the loss or decrease of OTC enzyme function, which hinders the ammonia conversion to urea, resulting in hyperammonemia and severe neurological dysfunction. Here, we studied one Chinese family of three generations who consecutively gave birth to two babies with OTCD. This study aims to explore the pathogenicity of two missense variants in the OTC gene and investigate the application of preimplantation genetic testing for monogenic (PGT-M) for a family troubled by Ornithine carbamoyltransferase deficiency (OTCD).MethodsThe retrospective method was used to classify the pathogenicity of two missense variants in the OTC gene in a family tortured by OTCD. Sanger sequencing was used to validate the variants in the OTC gene, and then the pathogenicity of variants was confirmed through family analysis and bioinformatics software. We used PGT-M to target the OTC gene and select a suitable embryo for transplantation. Prenatal diagnosis was recommended to confirm previous results using Sanger sequencing and karyotyping at an appropriate gestational stage. Tandem mass spectrometry (MS-MS) and gas chromatography-mass spectrometry (GC-MS) were used to detect fetal metabolism after birth. The number of the study cohort is ChiCTR2100053616.ResultsTwo missense variants, c.959G > C (p.Arg320Pro) and c.634G > A (p.Gly212Arg), were validated in the OTC gene in this family. According to the ACMG genetic variation classification criteria, the missense variant c.959G > C can be considered as “pathogenic”, and the missense variant c.634G > A can be regarded as “likely benign.” PGT-M identified a female embryo carrying the heterozygous variant c.959G > C (p.Arg320Pro), which was selected for transplantation. Prenatal diagnosis revealed the same variant in the fetus, and continued pregnancy was recommended. A female baby was born, and her blood amino acid testing and urine organic acid testing were regular. Follow-up was conducted after six months and indicated the girl was healthy.ConclusionOur research first validated the segregation of both c.959G > C and c.634G > A variants in the OTC gene in a Chinese OTCD family. Then, we classified variant c.959G > C as “pathogenic” and variant c.634G > A as “likely benign”, providing corresponding theoretical support for genetic counseling and fertility guidance in this family. PGT-M and prenatal diagnosis were recommended to help the couple receive a female baby successfully with a six-month follow-up.

Deciphering the roles of nitrogen source in sharping synchronous metabolic pathways of linear alkylbenzene sulfonate and nitrogen in a membrane biofilm for treating greywater

Nitrogen (N) source is an important factor affecting biological wastewater treatment. Although the oxygen-based membrane biofilm showed excellent greywater treatment performance, how N source impacts the synchronous removal of organics and N is still unclear. In this work, how N species (urea, nitrate and ammonia) affect synchronous metabolic pathways of organics and N were evaluated during greywater treatment in the membrane biofilm. Urea and ammonia achieved efficient chemical oxygen demand (>97.5%) and linear alkylbenzene sulfonate (LAS, >98.5%) removal, but nitrate enabled the maximum total N removal (80.8 ± 2.6%). The nitrate-added system had poor LAS removal ratio and high residual LAS, promoting the accumulation of effluent protein-like organics and fulvic acid matter. N source significantly induced bacterial community succession, and the increasing of corresponded functional flora can promote the transformation and utilization of microbial-mediated N. The nitrate system was more conducive to the accumulation of denitrification related microorganisms and enzymes, enabling the efficient N removal. Combining with high amount of ammonia monooxygenase that contributing to LAS and N co-metabolism, LAS mineralization related microbes and functional enzymes were generously accumulated in the urea and ammonia systems, which achieved the high efficiency of organics and LAS removal.