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                        "en": "Abstract Background Chitin, the main form of aminated polysaccharide in nature, is a biocompatible, polycationic, and antimicrobial biopolymer used extensively in industrial processes. Despite the abundance of chitin, applications thereof are hampered by difficulties in feedstock harvesting and limited structural versatility. To address these problems, we proposed a two-step cascade employing carbohydrate oxidoreductases and amine transaminases for plant polysaccharide aminations via one-pot reactions. Using a galactose oxidase from Fusarium graminearum for oxidation, this study compared the performance of CvATA (from Chromobacterium violaceum) and SpATA (from Silicibacter pomeroyi) on a range of oxidized carbohydrates with various structures and sizes. Using a rational enzyme engineering approach, four point mutations were introduced on the SpATA surface, and their effects on enzyme activity were evaluated. Results Herein, a quantitative colorimetric assay was developed to enable simple and accurate time-course measurement of the yield of transamination reactions. With higher operational stability, SpATA produced higher product yields in 36 h reactions despite its lower initial activity. Successful amination of oxidized galactomannan by SpATA was confirmed using a deuterium labeling method; higher aminated carbohydrate yields achieved with SpATA compared to CvATA were verified using HPLC and XPS. By balancing the oxidase and transaminase loadings, improved operating conditions were identified where the side product formation was largely suppressed without negatively impacting the product yield. SpATA mutants with multiple alanine substitutions besides E407A showed improved product yield. The E407A mutation reduced SpATA activity substantially, supporting its predicted role in maintaining the dimeric enzyme structure. Conclusions Using oxidase–amine transaminase cascades, the study demonstrated a fully enzymatic route to polysaccharide amination. Although the activity of SpATA may be further improved via enzyme engineering, the low operational stability of characterized amine transaminases, as a result of low retention of PMP cofactors, was identified as a key factor limiting the yield of the designed cascade. To increase the process feasibility, future efforts to engineer improved SpATA variants should focus on improving the cofactor affinity, and thus the operational stability of the enzyme. Graphical Abstract"
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                        "en": "Abstract Estradiol (E2) is a hormone that influences various aspects of women’s health. Beyond its reproductive functions, E2 impacts neurotransmitter systems such as dopamine (DA). Vertically aligned carbon nanofibers (VACNFs) have shown good sensitivity, selectivity against ascorbic acid (AA) and uric acid (UA), biocompatibility, and reduced fouling in DA sensing. In this study, we explore the use of Ti-Ni-CNF electrodes with CNFs grown for 5 min and 30 min for the direct electrochemical co-detection of E2 and DA. The longer growth time led to a 142% increase in average CNF length and a 36% larger electroactive surface area. In E2 detection, the electrodes demonstrate a wide linear range of 0.05–10 µM and sensitivity of 0.016 and 0.020 µA/µM for Ti-Ni-CNF-5 min and Ti-Ni-CNF-30 min, respectively. The sensor performance remains largely unaffected even in the presence of other steroid hormones such as progesterone and testosterone. Co-detection of equimolar E2 and DA shows promising peak separation of 0.34 ± 0.01 V and repeatability after 10 measurements. A notable improvement in the E2/DA peak current ratio, from 0.53 ± 0.07 to 0.81 ± 0.16, was achieved with the increased CNF length. Our results demonstrate the influence of adsorption sites in electrochemical detection, especially for analytes such as E2 and DA that both rely on adsorption for oxidation. While detecting small and fluctuating physiological concentrations remains a challenge, these findings can be used in choosing and fabricating electrode materials for more accurate and accessible continuous hormone measurements, including the possibility of multianalyte sensing platforms. Graphic abstract"
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