Enzyme and Microbial Technology最新文献

{"title":"Convenient preparation of indigo from the Ieaves of Baphicacanthus cusia(Nees) Bremek by enzymatic method and its MALDI-TOF-MS and UPLC-Q-TOF/MS analysis","authors":"HongXia Chen ,&nbsp;Hao Zhou ,&nbsp;Changwei Zhang ,&nbsp;Wenjun Li ,&nbsp;Xingying Xue ,&nbsp;ChengZhang Wang","doi":"10.1016/j.enzmictec.2024.110440","DOIUrl":"https://doi.org/10.1016/j.enzmictec.2024.110440","url":null,"abstract":"<div><p>The manufacturing of indigo naturalis requires prolonged leaf soaking and lime stirring; the resulting indigo purity is less than 3.00% and the yield of indigo (measured in stems and leaves weight) is less than 0.50%, making it unsuitable for use in industrial procedures like printing and dyeing. An enzymatic method of creating indigo without the requirement for lime was investigated in order to generate high purity indigo. Single factor tests were performed to optimize the enzymatic preparation conditions. The findings showed that 60 °C, pH 5.5, 200 mL of leaves extract containing 0.45 mg/mL indican, and a 4:1 ratio of the acidic cellulose (activity: 9000 U/mL, liquid) to indican were the ideal parameters for enzymatic preparation. The yield of indigo was 40.32%, and the contents of indigo and indirubin were 37.37% and 2.30%, respectively. MALDI-TOF-MS in positive ion mode and UPLC-Q-TOF-MS in both positive and negative ion modes were used to analyze indigo extracts from <em>Baphicacanthus cusia</em>(Nees) Bremek by enzymatic preparation. It has been discovered that 13 alkaloids, 5 organic acids, 3 terpenoids, 3 steroids, 2 flavones, and 7 other compounds are present in indigo extracts. The presence of the indigo, indirubin, isorhamnetin, tryptanthrin, indigodole B, and indigodole C determined by UPLC-Q-TOF-MS was verified by MALDI-TOF-MS analysis. The enzymatic preparation of indigo extracts kept the same chemical makeup as conventional indigo naturalis. Thermal analysis and SEM morphology were used to confirm that there was no lime in the indigo extract. During the enzymatic process, <em>Baphicacanthus cusia</em> (Nees) Bremek was employed more effectively, increasing the yield and purity of indigo.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"178 ","pages":"Article 110440"},"PeriodicalIF":3.4,"publicationDate":"2024-03-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140345254","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bioinspired mp20 mimicking uricase in ZIF-8: Metal ion dependent for controllable activity","authors":"Siti Fatimah Nur Abdul Aziz ,&nbsp;Abu Bakar Salleh ,&nbsp;Yahaya M. Normi ,&nbsp;Muhammad Alif Mohammad Latif ,&nbsp;Shahrul Ainliah Alang Ahmad","doi":"10.1016/j.enzmictec.2024.110439","DOIUrl":"10.1016/j.enzmictec.2024.110439","url":null,"abstract":"<div><p>Mini protein mimicking uricase (mp20) has shown significant potential as a replacement for natural enzymes in the development of uric acid biosensors. However, the design of mp20 has resulted to an inactive form of peptide, causing of loss their catalytic activity. Herein, this paper delineates the impact of various metal cofactors on the catalytic activity of mp20. The metal ion-binding site prediction and docking (MIB) web server was employed to identify the metal ion binding sites and their affinities towards mp20 residues. Among the tested metal ions, Cu²⁺ displayed the highest docking score, indicating its preference for interaction with Thr16 and Asp17 residues of mp20. To assess the catalytic activity of mp20 in the presence of metal ions, uric acid assays was monitored using a colorimetric method. The presence of Cu²⁺ in the assays promotes the activation of mp20, resulting in a color change based on quinoid production. Furthermore, the encapsulation of the mp20 within zeolitic imidazolate framework-8 (ZIF-8) notably improved the stability of the biomolecule. In comparison to the naked mp20, the encapsulated ZIFs biocomposite (mp20@ZIF-8) demonstrates superior stability, selectivity and sensitivity. ZIF’s porous shells provides excellent protection, broad detection (3–100 μM) with a low limit (4.4 μM), and optimal function across pH (3.4–11.4) and temperature (20–100°C) ranges. Cost-effective and stable mp20@ZIF-8 surpasses native uricase, marking a significant biosensor technology breakthrough. This integration of metal cofactor optimization and robust encapsulation sets new standards for biosensing applications.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"178 ","pages":"Article 110439"},"PeriodicalIF":3.4,"publicationDate":"2024-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140276200","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Inactivation of hydrogenase-3 leads to enhancement of 1,3-propanediol and 2,3-butanediol production by Klebsiella pneumoniae","authors":"Weiyan Jiang ,&nbsp;Yaoyu Cai ,&nbsp;Shaoqi Sun ,&nbsp;Wenqi Wang ,&nbsp;Marina Tišma ,&nbsp;Frank Baganz ,&nbsp;Jian Hao","doi":"10.1016/j.enzmictec.2024.110438","DOIUrl":"https://doi.org/10.1016/j.enzmictec.2024.110438","url":null,"abstract":"<div><p><em>Klebsiella pneumoniae</em> can use glucose or glycerol as carbon sources to produce 1,3-propanediol or 2,3-butanediol, respectively. In the metabolism of <em>Klebsiella pneumoniae</em>, hydrogenase-3 is responsible for H₂ production from formic acid, but it is not directly related to the synthesis pathways for 1,3-propanediol and 2,3-butanediol. In the first part of this research, <em>hycEFG</em>, which encodes subunits of the enzyme hydrogenase-3, was knocked out, so <em>K. pneumoniae</em> Δ<em>hycEFG</em> lost the ability to produce H₂ during cultivation using glycerol as a carbon source. As a consequence, the concentration of 1,3-propanediol increased and the substrate (glycerol) conversion ratio reached 0.587 mol/mol. Then, <em>K. pneumoniae</em> Δ<em>ldhA</em>Δ<em>hycEFG</em> was constructed to erase lactic acid synthesis which led to the further increase of 1,3-propanediol concentration. A substrate (glycerol) conversion ratio of 0.628 mol/mol in batch conditions was achieved, which was higher compared to the wild type strain (0.545 mol/mol). Furthermore, since <em>adhE</em> encodes an alcohol dehydrogenase that catalyzes ethanol production from acetaldehyde, <em>K. pneumoniae</em> Δ<em>ldhA</em>Δ<em>adhE</em>Δ<em>hycEFG</em> was constructed to prevent ethanol production. Contrary to expectations, this did not lead to a further increase, but to a decrease in 1,3-propanediol production. In the second part of this research, glucose was used as the carbon source to produce 2,3-butanediol. Knocking out <em>hycEFG</em> had distinct positive effect on 2,3-butanediol production. Especially in <em>K. pneumoniae ΔldhAΔadhEΔhycEFG</em>, a substrate (glucose) conversion ratio of 0.730 mol/mol was reached, which is higher compared to wild type strain (0.504 mol/mol). This work suggests that the inactivation of hydrogenase-3 may have a global effect on the metabolic regulation of <em>K. pneumoniae</em>, leading to the improvement of the production of two industrially important bulk chemicals, 1,3-propanediol and 2,3-butanediol.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110438"},"PeriodicalIF":3.4,"publicationDate":"2024-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140187802","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biosynthesis of mannose from glucose via constructing phosphorylation-dephosphorylation reactions in Escherichia coli","authors":"Yuyao Wang ,&nbsp;Enhui Chen ,&nbsp;Yanfei Wang ,&nbsp;Xinming Sun ,&nbsp;Qianzhen Dong ,&nbsp;Peng Chen ,&nbsp;Chenglin Zhang ,&nbsp;Jiangang Yang ,&nbsp;Yuanxia Sun","doi":"10.1016/j.enzmictec.2024.110427","DOIUrl":"https://doi.org/10.1016/j.enzmictec.2024.110427","url":null,"abstract":"<div><p><span>d</span>-mannose has been widely used in food, medicine, cosmetic, and food-additive industries. To date, chemical synthesis or enzymatic conversion approaches based on iso/epimerization reactions for <span>d</span>-mannose production suffered from low conversion rate due to the reaction equilibrium, necessitating intricate separation processes for obtaining pure products on an industrial scale. To circumvent this challenge, this study showcased a new approach for <span>d</span>-mannose synthesis from glucose through constructing a phosphorylation-dephosphorylation pathway in an engineered strain. Specifically, the gene encoding phosphofructokinase (PfkA) in glycolytic pathway was deleted in <em>Escherichia coli</em> to accumulate fructose-6-phosphate (F6P). Additionally, one endogenous phosphatase, YniC, with high specificity to mannose-6-phosphate, was identified. In Δ<em>pfkA</em> strain, a recombinant synthetic pathway based on mannose-6-phosphate isomerase and YniC was developed to direct F6P to mannose. The resulting strain successfully produced 25.2 g/L mannose from glucose with a high conversion rate of 63% after transformation for 48 h. This performance surpassed the 15% conversion rate observed with 2-epimerases. In conclusion, this study presents an efficient method for achieving high-yield mannose synthesis from cost-effective glucose.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110427"},"PeriodicalIF":3.4,"publicationDate":"2024-03-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140180367","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of sensitivity of Extended Gate Field Effect Transistor -biosensor based on V2O5/GOx for glucose detection","authors":"Alessandra Teixeira Felix ,&nbsp;Marcelo Mulato ,&nbsp;Elidia Maria Guerra","doi":"10.1016/j.enzmictec.2024.110428","DOIUrl":"10.1016/j.enzmictec.2024.110428","url":null,"abstract":"<div><p>The sensing modified electrode was prepared using glucose oxidase immobilized onto vanadium pentoxide xerogel with glass/FTO as support electrode to evaluate the possibility to construct a V₂O₅/GOx Extended Gate Field Effect Transistor biosensor. Previously, our studies exhibited a sensitivity of V₂O₅ of 58.1 mV/pH. The use of Nafion® onto V₂O₅/GOx caused a decrease of mass loss after several cycles compared to the modified electrode without Nafion® during the EQCM and cyclic voltammetrics studies. Electrical characterization of V₂O₅/GOx demonstrated a tendency to stability after 200 s as a function of applied current. In presence of glucose and in different pH, the current decreased when the glucose concentration increased due to the lower active sites of enzyme. After ten voltammetric cycles, the total charge tends to structural stability. In pH = 5.0, the modified electrode based on V₂O₅/GOx Extended Gate Field Effect Transistor presented more tendency to sensitivity in different concentration of glucose.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110428"},"PeriodicalIF":3.4,"publicationDate":"2024-03-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140279333","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Towards consolidated bioprocessing of biomass and plastic substrates for semi-synthetic production of bio-poly(ethylene furanoate) (PEF) polymer using omics-guided construction of artificial microbial consortia","authors":"Mohd Norfikri Omar ,&nbsp;Matthlessa Matthew Minggu ,&nbsp;Nor Azlan Nor Muhammad ,&nbsp;Peer Mohamed Abdul ,&nbsp;Ying Zhang ,&nbsp;Ahmad Bazli Ramzi","doi":"10.1016/j.enzmictec.2024.110429","DOIUrl":"10.1016/j.enzmictec.2024.110429","url":null,"abstract":"<div><p>Poly(ethylene furanoate) (PEF) plastic is a 100% renewable polyester that is currently being pursued for commercialization as the next-generation bio-based plastic. This is in line with growing demand for circular bioeconomy and new plastics economy that is aimed at minimizing plastic waste mismanagement and lowering carbon footprint of plastics. However, the current catalytic route for the synthesis of PEF is impeded with technical challenges including high cost of pretreatment and catalyst refurbishment. On the other hand, the semi-biosynthetic route of PEF plastic production is of increased biotechnological interest. In particular, the PEF monomers (Furan dicarboxylic acid and ethylene glycol) can be synthesized via microbial-based biorefinery and purified for subsequent catalyst-mediated polycondensation into PEF. Several bioengineering and bioprocessing issues such as efficient substrate utilization and pathway optimization need to be addressed prior to establishing industrial-scale production of the monomers. This review highlights current advances in semi-biosynthetic production of PEF monomers using consolidated waste biorefinery strategies, with an emphasis on the employment of omics-driven systems biology approaches in enzyme discovery and pathway construction. The roles of microbial protein transporters will be discussed, especially in terms of improving substrate uptake and utilization from lignocellulosic biomass, as well as from depolymerized plastic waste as potential bio-feedstock. The employment of artificial bioengineered microbial consortia will also be highlighted to provide streamlined systems and synthetic biology strategies for bio-based PEF monomer production using both plant biomass and plastic-derived substrates, which are important for circular and new plastics economy advances.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110429"},"PeriodicalIF":3.4,"publicationDate":"2024-03-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140203250","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization and application of active human α2,6-sialyltransferases ST6GalNAc V and ST6GalNAc VI recombined in Escherichia coli","authors":"Caixia Pei ,&nbsp;Xinlv Peng ,&nbsp;Yiran Wu ,&nbsp;Runmiao Jiao ,&nbsp;Tiehai Li ,&nbsp;Siming Jiao ,&nbsp;Lei Zhou ,&nbsp;Jianjun Li ,&nbsp;Yuguang Du ,&nbsp;Eika W. Qian","doi":"10.1016/j.enzmictec.2024.110426","DOIUrl":"https://doi.org/10.1016/j.enzmictec.2024.110426","url":null,"abstract":"<div><p>Eukaryotic sialyltransferases play key roles in many physiological and pathological events. The expression of active human recombinant sialyltransferases in bacteria is still challenging. In the current study, the genes encoding human <em>N</em>-acetylgalactosaminide α2,6-sialyltransferase V (hST6GalNAc V) and <em>N</em>-acetylgalactosaminide α2,6-sialyltransferase VI (hST6GalNAc VI) lacking the <em>N</em>-terminal transmembrane domains were cloned into the expression vectors, pET-32a and pET-22b, respectively. Soluble and active forms of recombinant hST6GalNAc V and hST6GalNAc VI when coexpressed with the chaperone plasmid pGro7 were successfully achieved in <em>Escherichia coli</em>. Further, lactose (Lac), Lacto-<em>N</em>-triose II (LNT II), lacto-<em>N</em>-tetraose (LNT), and sialyllacto-<em>N</em>-tetraose a (LSTa) were used as acceptor substrates to investigate their activities and substrate specificities. Unexpectedly, both can transfer sialic acid onto all those substrates. Compared with hST6GalNAc V expressed in the mammalian cells, the recombinant two α2,6-sialyltransferases in bacteria displayed flexible substrate specificities and lower enzymatic efficiency. In addition, an important human milk oligosaccharide disialyllacto-<em>N</em>-tetraose (DSLNT) can be synthesized by both human α2,6-sialyltransferases expressed in <em>E. coli</em> using LSTa as an acceptor substrate. To the best of our knowledge, these two active human α2,6-sialyltransferases enzymes were expressed in bacteria for the first time. They showed a high potential to be applied in biotechnology and investigating the molecular mechanisms of biological and pathological interactions related to sialylated glycoconjugates.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110426"},"PeriodicalIF":3.4,"publicationDate":"2024-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140145172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolism of phenolic compounds catalyzed by Tomato CYP736A61","authors":"Thanh Dat Mai ,&nbsp;Hyun Min Kim ,&nbsp;Seo Young Park ,&nbsp;Sang Hoon Ma ,&nbsp;Ju Hui Do ,&nbsp;Won Choi ,&nbsp;Hye Min Jang ,&nbsp;Hyeon Bae Hwang ,&nbsp;Eun Gyeong Song ,&nbsp;Jae Sung Shim ,&nbsp;Young Hee Joung","doi":"10.1016/j.enzmictec.2024.110425","DOIUrl":"https://doi.org/10.1016/j.enzmictec.2024.110425","url":null,"abstract":"<div><p>Cytochrome P450s (CYPs) regulate plant growth and stress responses by producing diverse primary and secondary metabolites. However, the function of many plant CYPs remains unknown because, despite their structural similarity, predicting the enzymatic activity of CYPs is difficult. In this study, one member of the CYP736A subfamily (CYP736A61) from tomatoes was isolated and characterized its enzymatic functions. CYP736A61 was successfully expressed in <em>Escherichia coli</em> through co-expression with molecular chaperones. The purified CYP736A61 showed hydroxylation activity toward 7-ethoxycoumarin, producing 7-hydroxycoumarin or 3-hydroxy 7-ethoxycoumarin. Further substrate screening revealed that dihydrochalcone and stilbene derivates (resveratrol and polydatin) are the substrates of CYP736A61. CYP736A61 also mediated the hydroxylation of resveratrol and polydatin, albeit with low activity. Importantly, CYP736A61 mediated the cleavage of resveratrol and polydatin as well as pinostilbene and pterostilbene. Interestingly, CY736A61 also converted phloretin to naringenin chalcone. These results suggest that CYP736A61 is a novel CYP enzyme with stilbene cleavage activity.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"176 ","pages":"Article 110425"},"PeriodicalIF":3.4,"publicationDate":"2024-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140103285","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Application of Thermomyces lanuginosus polygalacturonase produced in Komagataella phaffii in biomass hydrolysis and textile bioscouring","authors":"Luana Assis Serra ,&nbsp;Thais Demarchi Mendes ,&nbsp;Janice Lisboa De Marco ,&nbsp;João Ricardo Moreira de Almeida","doi":"10.1016/j.enzmictec.2024.110424","DOIUrl":"10.1016/j.enzmictec.2024.110424","url":null,"abstract":"<div><p>In this work, the polygalacturonase (TL-PG1) from the thermophilic fungus <em>Thermomyces lanuginosus</em> was heterologously produced for the first time in the yeast <em>Komagataella phaffii</em>. The TL-PG1 was successfully expressed under the control of the AOX1 promoter and sequentially purified by His-tag affinity. The purified recombinant pectinase exhibited an activity of 462.6 U/mL toward polygalacturonic acid under optimal conditions (pH 6 and 55 ˚C) with a 2.83 mg/mL and 0.063 μmol/minute for <em>K</em>_<em>m</em> and <em>V</em>_<em>max</em><em>,</em> respectively. When used as supplementation for biomass hydrolysis, TL-PG1 demonstrated synergy with the enzymatic cocktail Ctec3 to depolymerize orange citrus pulp, releasing 1.43 mg/mL of reducing sugar. In addition, TL-PG1 exhibited efficiency in fabric bioscouring, showing potential usage in the textile industry. Applying a protein dosage of 7 mg/mL, the time for the fabric to absorb water was 19.77 seconds (ten times faster than the control). Adding the surfactant Triton to the treatment allowed the reduction of the enzyme dosage by 50% and the water absorption time to 6.38 seconds. Altogether, this work describes a new versatile polygalacturonase from <em>T. lanuginosus</em> with the potential to be employed in the hydrolysis of lignocellulosic biomass and bioscouring.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"177 ","pages":"Article 110424"},"PeriodicalIF":3.4,"publicationDate":"2024-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140088312","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineered Zea mays phenylalanine ammonia-lyase for improve the catalytic efficiency of biosynthesis trans-cinnamic acid and p-coumaric acid","authors":"Jiangmei Zheng,&nbsp;Ruobin Sun,&nbsp;Dan Wu,&nbsp;Pengcheng Chen,&nbsp;Pu Zheng","doi":"10.1016/j.enzmictec.2024.110423","DOIUrl":"10.1016/j.enzmictec.2024.110423","url":null,"abstract":"<div><p>Phenylalanine ammonia-lyase (PAL) plays a pivotal role in the biosynthesis of phenylalanine. PAL from <em>Zea mays</em> (ZmPAL2) exhibits a bi-function of direct deamination of L-phenylalanine (L-Phe) or L-tyrosine(-L-Tyr) to form <em>trans</em>-cinnamic acid or <em>p</em>-coumaric acid. <em>trans</em>-Cinnamic acid and <em>p</em>-coumaric acid are mainly used in flavors and fragrances, food additives, pharmaceutical and other fields. Here, the Activity of ZmPAL2 toward L-Phe or L-Tyr was improved by using semi-rational and rational designs. The catalytic efficiency (<em>k</em>_cat/<em>K</em>_m) of mutant PT10 (V258I/I459V/Q484N) against L-Phe was 30.8 μM⁻¹ s⁻¹, a 4.5-fold increase compared to the parent, and the catalytic efficiency of mutant PA1 (F135H/I459L) to L-tyrosine exhibited 8.6 μM⁻¹ s⁻¹, which was 1.6-fold of the parent. The yield of <em>trans</em>-cinnamic acid in PT10 reached 30.75 g/L with a conversion rate of 98%. Meanwhile, PA1 converted L-Tyr to yield 3.12 g/L of <em>p</em>-coumaric acid with a conversion rate of 95%. Suggesting these two engineered ZmPAL2 to be valuable biocatalysts for the synthesis of <em>trans</em>-cinnamic acid and <em>p</em>-coumaric acid. In addition, MD simulations revealed that the underlying mechanisms of the increased catalytic efficiency of both mutant PT10 and PA1 are attributed to the substrate remaining stable within the pocket and closer to the catalytically active site. This also provides a new perspective on engineered PAL.</p></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"176 ","pages":"Article 110423"},"PeriodicalIF":3.4,"publicationDate":"2024-02-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139945470","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}