Applied Microbiology and Biotechnology最新文献

{"title":"Milk microbiome transplantation: recolonizing donor milk with mother's own milk microbiota.","authors":"Lisa F Stinson, Jie Ma, Ching Tat Lai, Alethea Rea, Sharon L Perrella, Donna T Geddes","doi":"10.1007/s00253-023-12965-8","DOIUrl":"10.1007/s00253-023-12965-8","url":null,"abstract":"<p><p>Donor human milk (DHM) provides myriad nutritional and immunological benefits for preterm and low birthweight infants. However, pasteurization leaves DHM devoid of potentially beneficial milk microbiota. In the present study, we performed milk microbiome transplantation from freshly collected mother's own milk (MOM) into pasteurized DHM. Small volumes of MOM (5%, 10%, or 30% v/v) were inoculated into pasteurized DHM and incubated at 37 °C for up to 8 h. Further, we compared microbiome recolonization in UV-C-treated and Holder-pasteurized DHM, as UV-C treatment has been shown to conserve important biochemical components of DHM that are lost during Holder pasteurization. Bacterial culture and viability-coupled metataxonomic sequencing were employed to assess the effectiveness of milk microbiome transplantation. Growth of transplanted MOM bacteria occurred rapidly in recolonized DHM samples; however, a greater level of growth was observed in Holder-pasteurized DHM compared to UV-C-treated DHM, potentially due to the conserved antimicrobial properties in UV-C-treated DHM. Viability-coupled metataxonomic analysis demonstrated similarity between recolonized DHM samples and fresh MOM samples, suggesting that the milk microbiome can be successfully transplanted into pasteurized DHM. These results highlight the potential of MOM microbiota transplantation to restore the microbial composition of UV-C-treated and Holder-pasteurized DHM and enhance the nutritional and immunological benefits of DHM for preterm and vulnerable infants. KEY POINTS: • Mother's own milk microbiome can be successfully transplanted into donor human milk. • Recolonization is equally successful in UV-C-treated and Holder-pasteurized milk. • Recolonization time should be restricted due to rapid bacterial growth.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"74"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10776751/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139401547","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Vaginal prevention of Candida albicans: synergistic effect of lactobacilli and mannan oligosaccharides (MOS).","authors":"Margarida Faustino, Joana Odila Pereira, Ana Margarida Pereira, Ana Sofia Oliveira, Carlos M H Ferreira, Carla F Pereira, Joana Durão, Manuela E Pintado, Ana P Carvalho","doi":"10.1007/s00253-023-12909-2","DOIUrl":"10.1007/s00253-023-12909-2","url":null,"abstract":"<p><p>Vulvovaginal candidiasis (VVC) affects approximately 30-50% of women at least once during their lifetime, causing uncomfortable symptoms and limitations in their daily quality of life. Antifungal therapy is not very effective, does not prevent recurrencies and usually causes side effects. Therefore, alternative therapies are urgently needed. The goal of this work was to investigate the potential benefits of using mannan oligosaccharides (MOS) extracts together with a Lactobacillus sp. pool, composed by the most significant species present in the vaginal environment, to prevent infections by Candida albicans. Microbial growth of isolated strains of the main vaginal lactobacilli and Candida strains was assessed in the presence of MOS, to screen their impact upon growth. A pool of the lactobacilli was then tested against C. albicans in competition and prophylaxis studies; bacterial and yeast cell numbers were quantified in specific time points, and the above-mentioned studies were assessed in simulated vaginal fluid (SVF). Finally, adhesion to vaginal epithelial cells (HeLa) was also evaluated, once again resorting to simultaneous exposure (competition) or prophylaxis assays, aiming to measure the effect of MOS presence in pathogen adherence. Results demonstrated that MOS extracts have potential to prevent vaginal candidiasis in synergy with vaginal lactobacilli, with improved results than those obtained when using lactobacilli alone. KEY POINTS: Potential benefits of MOS extracts with vaginal lactobacilli to prevent C. albicans infections. MOS impacts on growth of vaginal lactobacilli pool and C. albicans in SVF. MOS extracts in synergy with L. crispatus inhibit C. albicans adhesion in HeLa cells.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"73"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10776728/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139401552","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Adaptive laboratory evolution for improved tolerance of vitamin K in Bacillus subtilis.","authors":"Xiumin Ding, Zhiming Zheng, Genhai Zhao, Li Wang, Han Wang, Peng Wang","doi":"10.1007/s00253-023-12877-7","DOIUrl":"10.1007/s00253-023-12877-7","url":null,"abstract":"<p><p>Menaquinone-7 (MK-7), a subtype of vitamin K₂ (VK₂), assumes crucial roles in coagulation function, calcium homeostasis, and respiratory chain transmission. The production of MK-7 via microbial fermentation boasts mild technological conditions and high biocompatibility. Nevertheless, the redox activity of MK-7 imposes constraints on its excessive accumulation in microorganisms. To address this predicament, an adaptive laboratory evolution (ALE) protocol was implemented in Bacillus subtilis BS011, utilizing vitamin K₃ (VK₃) as a structural analog of MK-7. The resulting strain, BS012, exhibited heightened tolerance to high VK₃ concentrations and demonstrated substantial enhancements in biofilm formation and total antioxidant capacity (T-AOC) when compared to BS011. Furthermore, MK-7 production in BS012 exceeded that of BS011 by 76% and 22% under static and shaking cultivation conditions, respectively. The molecular basis underlying the superior performance of BS012 was elucidated through genome and transcriptome analyses, encompassing observations of alterations in cell morphology, variations in central carbon and nitrogen metabolism, spore formation, and antioxidant systems. In summation, ALE technology can notably enhance the tolerance of B. subtilis to VK and increase MK-7 production, thus offering a theoretical framework for the microbial fermentation production of other VK₂ subtypes. Additionally, the evolved strain BS012 can be developed for integration into probiotic formulations within the food industry to maintain intestinal flora homeostasis, mitigate osteoporosis risk, and reduce the incidence of cardiovascular disease. KEY POINTS: • Bacillus subtilis was evolved for improved vitamin K tolerance and menaquinone-7 (MK-7) production • Evolved strains formed wrinkled biofilms and elongated almost twofold in length • Evolved strains induced sporulation to improve tolerance when carbon was limited.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"75"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139401622","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":"Cleavage of cell junction proteins as a host invasion strategy in leptospirosis.","authors":"Preeti Kumari, Suhani Yadav, Sresha Sarkar, Padikara K Satheeshkumar","doi":"10.1007/s00253-023-12945-y","DOIUrl":"10.1007/s00253-023-12945-y","url":null,"abstract":"<p><p>Infection and invasion are the prerequisites for developing the disease symptoms in a host. While the probable mechanism of host invasion and pathogenesis is known in many pathogens, very little information is available on Leptospira invasion/pathogenesis. For causing systemic infection Leptospira must transmigrate across epithelial barriers, which is the most critical and challenging step. Extracellular and membrane-bound proteases play a crucial role in the invasion process. An extensive search for the proteins experimentally proven to be involved in the invasion process through cell junction cleavage in other pathogens has resulted in identifying 26 proteins. The similarity searches on the Leptospira genome for counterparts of these 26 pathogenesis-related proteins identified at least 12 probable coding sequences. The proteins were either extracellular or membrane-bound with a proteolytic domain to cleave the cell junction proteins. This review will emphasize our current understanding of the pathogenic aspects of host cell junction-pathogenic protein interactions involved in the invasion process. Further, potential candidate proteins with cell junction cleavage properties that may be exploited in the diagnostic/therapeutic aspects of leptospirosis will also be discussed. KEY POINTS: • The review focussed on the cell junction cleavage proteins in bacterial pathogenesis • Cell junction disruptors from Leptospira genome are identified using bioinformatics • The review provides insights into the therapeutic/diagnostic interventions possible.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"119"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10781872/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139416182","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Loop-mediated isothermal amplification linked a nanoparticles-based biosensor for detecting Epstein-Barr virus.","authors":"Xinggui Yang, Xiaoyan Zeng, Junfei Huang, Ludi Yang, Sha Mao, Xu Chen, Yu Wang, Xiaoyu Wei, Shijun Li","doi":"10.1007/s00253-023-12948-9","DOIUrl":"10.1007/s00253-023-12948-9","url":null,"abstract":"<p><p>Epstein-Barr virus (EBV) is a ubiquitous gamma herpesvirus that maintains a lifelong latent association with B lymphocytes. Here, a rapid and reliable diagnosis platform for detecting EBV infection, employing loop-mediated isothermal amplification (LAMP) combined with a gold nanoparticles-based lateral flow biosensors (AuNPs-LFB) (termed LAMP Amplification Mediated AuNPs-LFB Detection, LAMAD), was developed in the current study. A set of specific LAMP primers targeting the Epstein-Barr nuclear antigen (EBNA) leader protein (EBNA-LP) gene was designed and synthesized. Subsequently, these templates extracted from various pathogens and whole blood samples were used to optimize and evaluate the EBV-LAMAD assay. As a result, the limit of detection (LoD) of the EBV-LAMAD assay was 45 copies/reaction. The EBV-LAMAD assay can detect all representative EBV pathogens used in the study, and of note, no cross-reactions were observed with other non-EBV organisms. Moreover, the whole workflow of the EBV-LAMAD assay can be completed within 70 min, including rapid EBV template preparation, EBV-LAMP amplification, and AuNPs-LFB-mediated detection. Taken together, the EBV-LAMAD assay targeting the EBNA-LP gene is a rapid, simplified, sensitive, reliable, and easy-to-use detection protocol that can be used as a competitive potential diagnostic/screening tool for EBV infection in clinical settings, especially in basic laboratories in resource-limited regions. KEY POINTS: • A novel, simplified, and easy-to-use AuNPs-LFB biosensor was designed and prepared. • LAMP combined with an AuNPs-LFB targeting the novel EBNA-LP gene was established. • EBV-LAMAD is a rapid, sensitive, and reliable detection protocol for EBV infection.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"91"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10784390/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139429069","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel, robust peptidyl-lys metalloendopeptidase from Trametes coccinea recombinantly expressed in Komagataella phaffii.","authors":"Uzair Ahmed, Tobias Stadelmann, Daniel Heid, Berit Würtz, Jens Pfannstiel, Katrin Ochsenreither, Thomas Eisele","doi":"10.1007/s00253-023-12986-3","DOIUrl":"10.1007/s00253-023-12986-3","url":null,"abstract":"<p><p>A novel peptidyl-lys metalloendopeptidase (Tc-LysN) from Tramates coccinea was recombinantly expressed in Komagataella phaffii using the native pro-protein sequence. The peptidase was secreted into the culture broth as zymogen (~38 kDa) and mature enzyme (~19.8 kDa) simultaneously. The mature Tc-LysN was purified to homogeneity with a single step anion-exchange chromatography at pH 7.2. N-terminal sequencing using TMTpro Zero and mass spectrometry of the mature Tc-LysN indicated that the pro-peptide was cleaved between the amino acid positions 184 and 185 at the Kex2 cleavage site present in the native pro-protein sequence. The pH optimum of Tc-LysN was determined to be 5.0 while it maintained ≥60% activity between pH values 4.5-7.5 and ≥30% activity between pH values 8.5-10.0, indicating its broad applicability. The temperature maximum of Tc-LysN was determined to be 60 °C. After 18 h of incubation at 80 °C, Tc-LysN still retained ~20% activity. Organic solvents such as methanol and acetonitrile, at concentrations as high as 40% (v/v), were found to enhance Tc-LysN's activity up to ~100% and ~50%, respectively. Tc-LysN's thermostability, ability to withstand up to 8 M urea, tolerance to high concentrations of organic solvents, and an acidic pH optimum make it a viable candidate to be employed in proteomics workflows in which alkaline conditions might pose a challenge. The nano-LC-MS/MS analysis revealed bovine serum albumin (BSA)'s sequence coverage of 84% using Tc-LysN which was comparable to the sequence coverage of 90% by trypsin peptides. KEY POINTS: •A novel LysN from Trametes coccinea (Tc-LysN) was expressed in Komagataella phaffii and purified to homogeneity •Tc-LysN is thermostable, applicable over a broad pH range, and tolerates high concentrations of denaturants •Tc-LysN was successfully applied for protein digestion and mass spectrometry fingerprinting.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"103"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10787681/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139477440","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of fluorescence-linked immunosorbent assay for rapid detection of Staphylococcus aureus.","authors":"Joo-Kyung Kim, Hyun-Young Yun, Jae-Seok Kim, Wooseong Kim, Chang-Soo Lee, Byung-Gee Kim, Hee-Jin Jeong","doi":"10.1007/s00253-023-12836-2","DOIUrl":"10.1007/s00253-023-12836-2","url":null,"abstract":"<p><p>Staphylococcus aureus is a major pathogen that causes infections and life-threatening diseases. Although antibiotics, such as methicillin, have been used, methicillin-resistant S. aureus (MRSA) causes high morbidity and mortality rates, and conventional detection methods are difficult to be used because of time-consuming process. To control the spread of S. aureus, a development of a rapid and simple detection method is required. In this study, we generated a fluorescent anti-S. aureus antibody, and established a novel fluorescence-linked immunosorbent assay (FLISA)-based S. aureus detection method. The method showed high sensitivity and low limit of detection toward MRSA detection. The assay time for FLISA was 5 h, which was faster than that of conventional enzyme-linked immunosorbent assay (ELISA) or rapid ELISA. Moreover, the FLISA-based detection method was applied to diagnose clinically isolated MRSA samples that required only 5.3 h of preincubation. The FLISA method developed in this study can be widely applied as a useful tool for convenient S. aureus detection. KEY POINTS: • A fluorescence-linked immunosorbent assay-based S. aureus detection method • Simultaneous quantification of a maximum of 96 samples within 5 h • Application of the novel system to diagnosis clinical isolates.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"2"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139048254","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":"Research progress on carotenoid production by Rhodosporidium toruloides.","authors":"Zhuo-Ting Xie, Bing-Qian Mi, Yong-Jun Lu, Mou-Tong Chen, Zhi-Wei Ye","doi":"10.1007/s00253-023-12943-0","DOIUrl":"10.1007/s00253-023-12943-0","url":null,"abstract":"<p><p>Carotenoids are natural lipophilic pigments, which have been proven to provide significant health benefits to humans, relying on their capacity to efficiently scavenge singlet oxygen and peroxyl radicals as antioxidants. Strains belonging to the genus Rhodosporidium represent a heterogeneous group known for a number of phenotypic traits including accumulation of carotenoids and lipids and tolerance to heavy metals and oxidative stress. As a representative of these yeasts, Rhodosporidium toruloides naturally produces carotenoids with high antioxidant activity and grows on a wide variety of carbon sources. As a result, R. toruloides is a promising host for the efficient production of more value-added lipophilic compound carotenoids, e.g., torulene and torularhodin. This review provides a comprehensive summary of the research progress on carotenoid biosynthesis in R. toruloides, focusing on the understanding of biosynthetic pathways and the regulation of key enzymes and genes involved in the process. Moreover, the relationship between the accumulation of carotenoids and lipid biosynthesis, as well as the stress from diverse abiotic factors, has also been discussed for the first time. Finally, several feasible strategies have been proposed to promote carotenoid production by R. toruloides. It is possible that R. toruloides may become a critical strain in the production of carotenoids or high-value terpenoids by genetic technologies and optimal fermentation processes. KEY POINTS: • Biosynthetic pathway and its regulation of carotenoids in Rhodosporidium toruloides were concluded • Stimulation of abiotic factors for carotenoid biosynthesis in R. toruloides was summarized • Feasible strategies for increasing carotenoid production by R. toruloides were proposed.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"7"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139085672","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":"Development of multiplex allele-specific RT-qPCR assays for differentiation of SARS-CoV-2 Omicron subvariants.","authors":"Jianguo Li, Ruiling Cheng, Zixin Bian, Jiahui Niu, Juan Xia, Guoli Mao, Hulong Liu, Changxin Wu, Chunyan Hao","doi":"10.1007/s00253-023-12941-2","DOIUrl":"10.1007/s00253-023-12941-2","url":null,"abstract":"<p><p>Quick differentiation of current circulating variants and the emerging recombinant variants of SARS-CoV-2 is essential to monitor their transmissions. However, the widely applied gene sequencing method is time-consuming and costly especially when facing recombinant variants, because a large part or whole genome sequencing is required. Allele-specific reverse transcriptase real time RT-PCR (RT-qPCR) represents a quick and cost-effective method for SNP (single nucleotide polymorphism) genotyping and has been successfully applied for SARS-CoV-2 variant screening. In the present study, we developed a panel of 5 multiplex allele-specific RT-qPCR assays targeting 20 key mutations for quick differentiation of the Omicron subvariants (BA.1 to BA.5 and their descendants) and the recombinant variants (XBB.1 and XBB.1.5). Two parallel multiplex RT-qPCR reactions were designed to separately target the prototype allele and the mutated allele of each mutation in the allele-specific RT-qPCR assay. Optimal annealing temperatures, primer and probe dosage, and time for annealing/extension for each reaction were determined by multi-factor and multi-level orthogonal test. The variation of Cp (crossing point) values (ΔCp) between the two multiplex RT-qPCR reactions was applied to determine if a mutation occurs or not. SARS-CoV-2 subvariants and related recombinant variants were differentiated by their unique mutation patterns. The developed multiplex allele-specific RT-qPCR assays exhibited excellent analytical sensitivities (with limits of detection (LoDs) of 1.47-18.52 copies per reaction), wide linear detection ranges (10⁹-10⁰ copies per reaction), good amplification efficiencies (88.25 to 110.68%), excellent reproducibility (coefficient of variations (CVs) < 5% in both intra-assay and inter-assay tests), and good clinical performances (99.5-100% consistencies with Sanger sequencing). The developed multiplex allele-specific RT-qPCR assays in the present study provide an alternative tool for quick differentiation of the SARS-CoV-2 Omicron subvariants and their recombinant variants. KEY POINTS: • A panel of five multiplex allele-specific RT-qPCR assays for quick differentiation of 11 SARS-CoV-2 Omicron subvariants (BA.1, BA.2, BA.4, BA.5, and their descendants) and 2 recombinant variants (XBB.1 and XBB.1.5). • The developed assays exhibited good analytical sensitivities and reproducibility, wide linear detection ranges, and good clinical performances, providing an alternative tool for quick differentiation of the SARS-CoV-2 Omicron subvariants and their recombinant variants.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"35"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139110704","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":"Discovery and biochemical characterization of thermostable glycerol oxidases.","authors":"Lars L Santema, Laura Rotilio, Ruite Xiang, Gwen Tjallinks, Victor Guallar, Andrea Mattevi, Marco W Fraaije","doi":"10.1007/s00253-023-12883-9","DOIUrl":"10.1007/s00253-023-12883-9","url":null,"abstract":"<p><p>Alditol oxidases are promising tools for the biocatalytic oxidation of glycerol to more valuable chemicals. By integrating in silico bioprospecting with cell-free protein synthesis and activity screening, an effective pipeline was developed to rapidly identify enzymes that are active on glycerol. Three thermostable alditol oxidases from Actinobacteria Bacterium, Streptomyces thermoviolaceus, and Thermostaphylospora chromogena active on glycerol were discovered. The characterization of these three flavoenzymes demonstrated their glycerol oxidation activities, preference for alkaline conditions, and excellent thermostabilities with melting temperatures higher than 75 °C. Structural elucidation of the alditol oxidase from Actinobacteria Bacterium highlighted a constellation of side chains that engage the substrate through several hydrogen bonds, a histidine residue covalently bound to the FAD prosthetic group, and a tunnel leading to the active site. Upon computational simulations of substrate binding, a double mutant targeting a residue pair at the tunnel entrance was created and found to display an improved thermal stability and catalytic efficiency for glycerol oxidation. The hereby described alditol oxidases form a valuable panel of oxidative biocatalysts that can perform regioselective oxidation of glycerol and other polyols. KEY POINTS: • Rapid pipeline designed to identify putative oxidases • Biochemical and structural characterization of alditol oxidases • Glycerol oxidation to more valuable derivatives.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"108 1","pages":"61"},"PeriodicalIF":5.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10771423/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139110705","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}