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Adipic acid is an industrially important chemical, but the current approach to synthesize it can be of serious pollution to the environment. Rencently, bio-based production of adipic acid has significantly advanced with the development of metabolic engineering and synthetic biology. However, genetic heterogeneity-caused decrease of product titer has largely limited the industrialization of chemicals like adipic acid. Therefore, in the attempt to overcome this challenge, we constitutively expressed the reverse adipate degradation pathway, designed and optimized an adipic acid biosensor, and established a high-throughput screening platform to screen for high-performance strains based on the optimized biosensor. Using this platform, we successfully screened a strain with an adipic acid titer of 188.08 mg·L−1. Coupling the screening platform with fermentation optimization, the titer of adipic acid reached 531.88 mg·L−1 under shake flask fermentation, which achieved an 18.78-fold improvement comparing to the initial strain. Scale-up fermentation in a 5-L fermenter utilizing the screened high-performance strain was eventually conducted, in which the adipic acid titer reached 3.62 g·L−1. Overall, strategies developed in this study proved to be a potentially efficient method in reducing the genetic heterogeneity and was expected to provide guidance in helping to build a more efficient industrial screening process.Key points• Developed a fine-tuned adipic acid biosensor.• Established a high-throughput screening platform to screen high-performance strains.• The titer of adipic acid reached 3.62 g·L−1 in a 5-L fermenter.

Many new active pharmaceutical ingredients (APIs) demonstrate high hydrophobicity and low water‐solubility issues. In this regard, polymeric nanoparticles (NPs) have been extensively used as drug delivery carriers for the encapsulation of such APIs. One commonly used polymer is polyethylene glycol (PEG), owing to its biocompatibility, high water solubility, and capacity to prolong the drug residence time. However, concerns have arisen regarding PEG's immunogenicity and limited biodegradability. In addition, inherent limitations, including limited chemical handles can restrict PEG's effectiveness in physiological conditions. For this reason, in the present study, we combine the advantages offered by PEG with the use of an enzymatic synthetic route to produce novel PEGylated polyesters. Furthermore, it has been proven that incorporation of hydrophobic diols into the PEGylated backbone influences NPs formation, stability, and drug encapsulation, despite high chemical similarity. As a preliminary result, samples containing PEG and 1,6‐hexanediol in a 50 : 50 ratio (PEGA‐Hex 50 %) and PEG and 2‐hydroxyethyl disulfide in a 50 : 50 ratio (PEGA‐SS 50 %) have proved to be the most promising candidates in this small library analysed. Both samples exhibited sufficient NPs stability, biocompatibility, and superior encapsulation efficiency compared to the other variants. This study explores a one‐pot, one‐step enzymatic polycondensation process to create amphiphilic PEG‐adipate variants with improved physicochemical properties that can self‐assemble into nanoparticles. By varying hydrophobic and functionalised diols, the impact on nanoparticle formation, stability, and drug encapsulation is examined. The results highlight the potential of these PEGylated polymers for sustainable nano‐drug delivery systems.

Poly(butylene adipate-co-terephthalate) (PBAT), a polyester made of terephthalic acid (TPA), 1,4-butanediol, and adipic acid, is extensively utilized in plastic production and has accumulated globally as environmental waste. Biodegradation is an attractive strategy to manage PBAT, but an effective PBAT-degrading enzyme is required. Here, we demonstrate that cutinases are highly potent enzymes that can completely decompose PBAT films in 48 h. We further show that the engineered cutinases, by applying a double mutation strategy to render a more flexible substrate-binding pocket exhibit higher decomposition rates. Notably, these variants produce TPA as a major end-product, which is beneficial feature for the future recycling economy. The crystal structures of wild type and double mutation of a cutinase from Thermobifida fusca in complex with a substrate analogue are also solved, elucidating their substrate-binding modes. These structural and biochemical analyses enable us to propose the mechanism of cutinase-mediated PBAT degradation. Bio-degradation of poly(butylene adipate-co-terephthalate) is an attractive tactic but requires an effective hydrolytic enzyme. Here, the authors demonstrate that cutinases are highly potent PBAT-decomposing enzymes and their mechanism is proposed based on substrate-binding mode.

With the issue of plastic waste persisting and the need for more sustainable solutions to the ever‐increasing demand for lightweight and durable plastic products, this review has become imminent and compelling. Poly (butylene adipate‐co‐terephthalate) (PBAT) is a biodegradable polymer with exceptional film‐forming ability resembling those of low‐density polyethylene. PBAT has a huge advantage for packaging applications due to its remarkably high elongation at break, giving it a good processing window for its application in packaging. However, certain defiant intrinsic properties stand in the way of its full commercialization. The development of blends and biocomposites of PBAT has, therefore, become imperative for complementing its properties and producing a superior material. This paper focuses on the recent developments in preparing PBAT‐based blends and biocomposites with superior mechanical, barrier, and antimicrobial properties and, most importantly, has also investigated how the development of these blends and biocomposites impacts the biodegradation rate of PBAT. It also highlights the possible synthesis of bio‐based PBAT and the commercialization, market trends, and prospects of PBAT‐based materials for flexible, rigid packaging, and other industrial applications compared with biodegradable alternatives.

Plastic food contact materials (FCM)-based products were widely used in everyday life. These products were normally imposed to strict regulations in order to pass the enforcement tests of compliance as a prefix condition. However, even in these \"qualified\" materials, unknown chemical substances, not involving in legislation lists, could migrate from FCM. In this perspective, the present work aims to thoroughly analyze by means of Gas Chromatography-Mass Spectrometry (GC-MS) the different substances/migrants in 120 qualified FCM plastic products. Unexpectedly, among the identified compounds (nearly 100), only 13% was included in the permitted list of Commission Regulation EU No 10/2011. All the identified compounds were classified into 11 categories according to their chemical structure and the FCM type, whereas toxicology data were in addition analyzed. Each plastic type exhibited different preferences of chemical migrants. Fortunately, most of the compounds identified were of low toxicity, and only 4 chemicals were included in priority lists and previous literature reports as potential risk factors. Subsequently, the accurate amount of these 4 chemicals was determined. The amount of Bis(2-ethylhexyl) adipate (DEHA) and Bis(2-ethylhexyl) phthalate (DEHP) were lower than the SML in Commission Regulation EU No 10/2011, and that of stearamide was under the recommended use quantity. The 2,4-di-tert-butylphenol (2,4-DTBP) was widely exist in the investigated FCM products. Among them, the highest level is obtained in polypropylene/low density polyethylene (BOPP/LDPE) materials, up to 45.568±31.513 mg/kg. In summary, a panel of unlisted chemical migrants were discovered and identified by GS-MS screening. The results implied that plastic FCMs were not so \"inert\" as they usually considered, and further safety evaluation should be performed toward the complete identification of new substances in FCM products.

Measuring pulmonary nodules accurately can help the early diagnosis of lung cancer, which can increase the survival rate among patients. Numerous techniques for lung nodule segmentation have been developed; however, most of them either rely on the 3D volumetric region of interest (VOI) input by radiologists or use the 2D fixed region of interest (ROI) for all the slices of computed tomography (CT) scan. These methods only consider the presence of nodules within the given VOI, which limits the networks’ ability to detect nodules outside the VOI and can also encompass unnecessary structures in the VOI, leading to potentially inaccurate segmentation. In this work, we propose a novel approach for 3D lung nodule segmentation that utilizes the 2D region of interest (ROI) inputted from a radiologist or computer-aided detection (CADe) system. Concretely, we developed a two-stage lung nodule segmentation technique. Firstly, we designed a dual-encoder-based hard attention network (DEHA-Net) in which the full axial slice of thoracic computed tomography (CT) scan, along with an ROI mask, were considered as input to segment the lung nodule in the given slice. The output of DEHA-Net, the segmentation mask of the lung nodule, was inputted to the adaptive region of interest (A-ROI) algorithm to automatically generate the ROI masks for the surrounding slices, which eliminated the need for any further inputs from radiologists. After extracting the segmentation along the axial axis, at the second stage, we further investigated the lung nodule along sagittal and coronal views by employing DEHA-Net. All the estimated masks were inputted into the consensus module to obtain the final volumetric segmentation of the nodule. The proposed scheme was rigorously evaluated on the lung image database consortium and image database resource initiative (LIDC/IDRI) dataset, and an extensive analysis of the results was performed. The quantitative analysis showed that the proposed method not only improved the existing state-of-the-art methods in terms of dice score but also showed significant robustness against different types, shapes, and dimensions of the lung nodules. The proposed framework achieved the average dice score, sensitivity, and positive predictive value of 87.91%, 90.84%, and 89.56%, respectively.

Arthrobacter sp. strain KI72 grows on a 6-aminohexanoate oligomer, which is a by-product of nylon-6 manufacturing, as a sole source of carbon and nitrogen. We cloned the two genes, nylD1 and nylE1, responsible for 6-aminohexanoate metabolism on the basis of the draft genomic DNA sequence of strain KI72. We amplified the DNA fragments that encode these genes by polymerase chain reaction using a synthetic primer DNA homologous to the 4-aminobutyrate metabolic enzymes. We inserted the amplified DNA fragments into the expression vector pColdI in Escherichia coli, purified the His-tagged enzymes to homogeneity, and performed biochemical studies. We confirmed that 6-aminohexanoate aminotransferase (NylD1) catalyzes the reaction of 6-aminohexanoate to adipate semialdehyde using α-ketoglutarate, pyruvate, and glyoxylate as amino acceptors, generating glutamate, alanine, and glycine, respectively. The reaction requires pyridoxal phosphate (PLP) as a cofactor. For further metabolism, adipate semialdehyde dehydrogenase (NylE1) catalyzes the oxidative reaction of adipate semialdehyde to adipate using NADP+ as a cofactor. Phylogenic analysis revealed that NylD1 should be placed in a branch of the PLP-dependent aminotransferase sub III, while NylE1 should be in a branch of the aldehyde dehydrogenase superfamily. In addition, we established a NylD1/NylE1 coupled system to quantify the aminotransferase activity and to enable the conversion of 6-aminohexaoate to adipate via adipate semialdehyde with a yield of > 90%. In the present study, we demonstrate that 6-aminohexanoate produced from polymeric nylon-6 and nylon oligomers (i.e., a mixture of 6-aminohexaoate oligomers) by nylon hydrolase (NylC) and 6-aminohexanoate dimer hydrolase (NylB) reactions are sequentially converted to adipate by metabolic engineering technology.

The biological significance of the DHTKD1- encoded 2-oxoadipate dehydrogenase (OADH) remains obscure due to its catalytic redundancy with the ubiquitous OGDH -encoded 2-oxoglutarate dehydrogenase (OGDH). In this work, metabolic contributions of OADH and OGDH are discriminated by exposure of cells/tissues with different DHTKD1 expression to the synthesized phosphonate analogues of homologous 2-oxodicarboxylates. The saccharopine pathway intermediates and phosphorylated sugars are abundant when cellular expressions of DHTKD1 and OGDH are comparable, while nicotinate and non-phosphorylated sugars are when DHTKD1 expression is order(s) of magnitude lower than that of OGDH . Using succinyl, glutaryl and adipoyl phosphonates on the enzyme preparations from tissues with varied DHTKD1 expression reveals the contributions of OADH and OGDH to oxidation of 2-oxoadipate and 2-oxoglutarate in vitro . In the phosphonates-treated cells with the high and low DHTKD1 expression, adipate or glutarate, correspondingly, are the most affected metabolites. The marker of fatty acid β-oxidation, adipate, is mostly decreased by the shorter, OGDH-preferring, phosphonate, in agreement with the known OGDH dependence of β-oxidation. The longest, OADH-preferring, phosphonate mostly affects the glutarate level. Coupled decreases in sugars and nicotinate upon the OADH inhibition link the perturbation in glucose homeostasis, known in OADH mutants, to the nicotinate-dependent NAD metabolism.

Purpose To understand the role of the mitochondrial oxodicarboxylate carrier (SLC25A21) in the development of spinal muscular atrophy-like disease. Methods We identified a novel pathogenic variant in a patient by whole-exome sequencing. The pathogenicity of the mutation was studied by transport assays, computer modeling, followed by targeted metabolic testing and in vitro studies in human fibroblasts and neurons. Results The patient carries a homozygous pathogenic variant c.695A>G; p.(Lys232Arg) in the SLC25A21 gene, encoding the mitochondrial oxodicarboxylate carrier, and developed spinal muscular atrophy and mitochondrial myopathy. Transport assays show that the mutation renders SLC25A21 dysfunctional and 2-oxoadipate cannot be imported into the mitochondrial matrix. Computer models of central metabolism predicted that impaired transport of oxodicarboxylate disrupts the pathways of lysine and tryptophan degradation, and causes accumulation of 2-oxoadipate, pipecolic acid, and quinolinic acid, which was confirmed in the patient’s urine by targeted metabolomics. Exposure to 2-oxoadipate and quinolinic acid decreased the level of mitochondrial complexes in neuronal cells (SH-SY5Y) and induced apoptosis. Conclusion Mitochondrial oxodicarboxylate carrier deficiency leads to mitochondrial dysfunction and the accumulation of oxoadipate and quinolinic acid, which in turn cause toxicity in spinal motor neurons leading to spinal muscular atrophy–like disease.

The β-ketoadipate pathway (β-KAP) is an enzyme-mediated aryl-ring degradation sequence employed by a wide selection of soil bacteria and fungi to reconcile the conversion of many hazardous aromatic pollutants into benign metabolites of the tricarboxylic acid cycle (TCA), lipogenesis, and other anabolic processes. Recently determined catabolic sequences that incorporate the β-KAP allow thorough mineralization of toxic priority pollutants including hazardous nitrophenols, organophosphates, and polychlorinated arenes and hydrocarbons. Novel investigations have applied the β-KAP via oleaginous microorganisms to convert aromatic lignocellulosic waste into bio-oils that are suitable for biodiesel applications. These newly elucidated catabolic pathways and applications, reviewed here, provide exciting and unparalleled biotechnological opportunities for the future.