Biotechnology and Bioengineering最新文献

Metabolic Engineering of Nonmodel Yeast Issatchenkia orientalis SD108 for 5-Aminolevulinic Acid Production. 生产 5-氨基乙酰丙酸的非模式酵母 Issatchenkia orientalis SD108 的代谢工程。

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-11-06 DOI: 10.1002/bit.28877

Shih-I Tan, I-Son Ng, Huimin Zhao

{"title":"Metabolic Engineering of Nonmodel Yeast Issatchenkia orientalis SD108 for 5-Aminolevulinic Acid Production.","authors":"Shih-I Tan, I-Son Ng, Huimin Zhao","doi":"10.1002/bit.28877","DOIUrl":"10.1002/bit.28877","url":null,"abstract":"Biological production of 5-aminolevulinic acid (5-ALA) has received growing attention over the years. However, there is the tradeoff between 5-ALA biosynthesis and cell growth because the fermentation broth will become acidic due to the production of 5-ALA. To address this limitation, we engineered an acid-tolerant yeast, Issatchenkia orientalis SD108, for 5-ALA production. We first discovered that the cell growth rate of I. orientalis SD108 was boosted by 5-ALA and its endogenous ALA synthetase (ALAS) showed higher activity than those homologs from other yeasts. The titer of 5-ALA was improved from 28 mg/L to 120-, 150-, and 300 mg/L, by optimizing plasmid design, overexpressing a transporter, and increasing gene copy number, respectively. After redirecting the metabolic flux using the pyruvate decarboxylase (PDC) knockout strain (SD108ΔPDC) and culturing with urea, we increased the titer of 5-ALA to 510 mg/L, a 13-fold enhancement, proving the importance of the newly identified IoALAS with higher activity and the strategic selection of nitrogen sources for knockout strains. This study demonstrates the acid-tolerant I. orientalis SD108ΔPDC has a high potential for 5-ALA production at a large scale in the future.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142589456","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Oxygen Consumption in Filamentous Pellets of Aspergillus niger: Microelectrode Measurements and Modeling. 黑曲霉丝状颗粒的耗氧量：微电极测量与建模

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-11-04 DOI: 10.1002/bit.28874

Charlotte Deffur, Anna Dinius, Julian Pagel, Henri Müller, Stefan Schmideder, Heiko Briesen, Rainer Krull

{"title":"Oxygen Consumption in Filamentous Pellets of Aspergillus niger: Microelectrode Measurements and Modeling.","authors":"Charlotte Deffur, Anna Dinius, Julian Pagel, Henri Müller, Stefan Schmideder, Heiko Briesen, Rainer Krull","doi":"10.1002/bit.28874","DOIUrl":"https://doi.org/10.1002/bit.28874","url":null,"abstract":"Filamentous fungi cultivated as biopellets are well established in biotechnology industries. A distinctive feature of filamentous fungi is that hyphal growth and fungal morphology affect product titers and require tailored process conditions. Within the pellet, mass transfer, substrate consumption, and biomass formation are intricately linked to the local hyphal fraction and pellet size. This study combined oxygen concentration measurements with microelectrode profiling and three-dimensional X-ray microtomography measurements of the same fungal pellets for the first time. This allowed for the precise correlation of micromorphological information with local oxygen concentrations of two Aspergillus niger strains (hyperbranching and regular branching). The generated results showed that the identified oxygen-penetrated outer pellet regions exhibited a depth of 90-290 µm, strain-specific, with the active part percentage in the pellet ranging from 18% to 69%, without any difference between strains. Using a 1D continuum diffusion consumption model, the oxygen concentration in the pellets was computed depending on the local hyphal fraction. The best simulation results were achieved by individually estimating the oxygen-related biomass yield coefficient of the consumption term within each examined pellet, with an average estimated value of 1.95 (± 0.72) kg biomass per kg oxygen. The study lays the foundation for understanding oxygen supply in fungal pellets and optimizing processes and pellet morphologies accordingly.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142575324","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Urea-Loaded PLGA Microspheres as Chemotaxis Stimulants for Helicobacter pylori. 作为幽门螺旋杆菌趋化刺激剂的尿素负载聚乳酸聚乳酸（PLGA）微球

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-11-03 DOI: 10.1002/bit.28870

Prasanth Shanmughan, Pravin Subrahmaniyan, Dhruv Bhatnagar, Srinithi Ranganathan, Pushkar P Lele

{"title":"Urea-Loaded PLGA Microspheres as Chemotaxis Stimulants for Helicobacter pylori.","authors":"Prasanth Shanmughan, Pravin Subrahmaniyan, Dhruv Bhatnagar, Srinithi Ranganathan, Pushkar P Lele","doi":"10.1002/bit.28870","DOIUrl":"https://doi.org/10.1002/bit.28870","url":null,"abstract":"Helicobacter pylori cells undergo chemotaxis toward several small molecules, called chemo-attractants, including urea produced by the epithelial cells of the stomach. The biophysical mechanisms of chemotaxis are not well understood in H. pylori. Here, we developed point sources of urea by encapsulating it in Poly(lactic-co-glycolic acid) or PLGA microbeads for H. pylori chemotaxis studies. Microscopy and Dynamic Light Scattering characterization indicated that the PLGA particles had an average diameter of < 0.8 μm. The particles were relatively stable and had a net negative surface charge. Absorbance measurements indicated that the beads released ~70% of the urea over a 2-week period, with most of the release occurring within the first 24-h period. Varying pH (2.0-7.0) had little effect on the rate of urea release. A diffusion model predicted that such beads could generate sufficient urea gradients to chemotactically attract H. pylori cells. Single-bead single-cell chemotaxis assays confirmed the predictions, revealing that H. pylori continued to be attracted to beads even after most of the urea had been released in the first 24 h. Our work highlights a novel use of PLGA microbeads as delivery vehicles for stimulating a chemotaxis response in H. pylori, with potential applications in bacterial eradication strategies.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-11-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142567101","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Design an Energy-Conserving Pathway for Efficient Biosynthesis of 1,5-Pentanediol and 5-Amino-1-Pentanol. 为 1,5-戊二醇和 5-氨基-1-戊醇的高效生物合成设计节能途径。

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-31 DOI: 10.1002/bit.28875

Lin Ma, Chong Xie, Yu Zhang, Wenna Li, Ning An, Xiaolin Shen, Jia Wang, Xinxiao Sun, Qipeng Yuan

引用次数: 0

Protein Scaffold-Mediated Multi-Enzyme Self-Assembly and Ordered Co-Immobilization of Flavin-Dependent Halogenase-Coenzyme Cycle System for Efficient Biosynthesis of 6-Cl-L-Trp. 蛋白支架介导的多酶自组装和黄素依赖性卤化酶-辅酶循环系统的有序协同固定，以实现 6-Cl-L-Trp 的高效生物合成。

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-28 DOI: 10.1002/bit.28871

Han-Yu Liu, Pan Ning, Feng Qian, Yao-Wu Wang, Hai-Min Zhang, Pu Wang

{"title":"Protein Scaffold-Mediated Multi-Enzyme Self-Assembly and Ordered Co-Immobilization of Flavin-Dependent Halogenase-Coenzyme Cycle System for Efficient Biosynthesis of 6-Cl-L-Trp.","authors":"Han-Yu Liu, Pan Ning, Feng Qian, Yao-Wu Wang, Hai-Min Zhang, Pu Wang","doi":"10.1002/bit.28871","DOIUrl":"https://doi.org/10.1002/bit.28871","url":null,"abstract":"Flavin-dependent halogenase (FDH) is highly prized in pharmaceutical and chemical industries for its exceptional capacity to produce halogenated aromatic compounds with precise regioselectivity. This study has devised a multi-enzyme self-assembly strategy to construct an effective and reliable in vitro coenzyme cycling system tailored for FDHs. Initially, tri-enzyme self-assembling nanoclusters (TESNCs) were developed, comprising glucose dehydrogenase (GDH), flavin reductase (FR) and FDH. The TESNCs exhibited enhanced thermal stability and conversion efficiency compared to free triple enzyme mixtures during the conversion of L-Trp to 6-Cl-L-Trp, resulting in a 2.1-fold increase in yield. Subsequently, an ordered co-immobilization of GDH, FR, and FDH was established, further amplifying the stability and catalytic efficiency of the FDH coenzyme cycle system. Compared to the free TESNCs, the immobilized TESNCs demonstrated a 4.2-fold increase in catalytic efficiency in a 5 mL reaction system. This research provides an effective strategy for developing a robust and efficient coenzyme recycling system for FDHs.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142521041","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Defining Golden Batches in Biomanufacturing Processes From Internal Metabolic Activity to Detect Process Changes That May Affect Product Quality. 从内部代谢活动定义生物制造过程中的黄金批次，以检测可能影响产品质量的过程变化。

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-27 DOI: 10.1002/bit.28873

Xin Bush, Erica J Fratz-Berilla, Casey L Kohnhorst, Roberta King, Cyrus Agarabi, David N Powers, Nicholas Trunfio

{"title":"Defining Golden Batches in Biomanufacturing Processes From Internal Metabolic Activity to Detect Process Changes That May Affect Product Quality.","authors":"Xin Bush, Erica J Fratz-Berilla, Casey L Kohnhorst, Roberta King, Cyrus Agarabi, David N Powers, Nicholas Trunfio","doi":"10.1002/bit.28873","DOIUrl":"https://doi.org/10.1002/bit.28873","url":null,"abstract":"Cellular metabolism plays a role in the observed variability of a drug substance's Critical Quality Attributes (CQAs) made by biomanufacturing processes. Therefore, here we describe a new approach for monitoring biomanufacturing processes that measures a set of metabolic reaction rates (named Critical Metabolic Parameters (CMP) in addition to the macroscopic process conditions currently being used as Critical Process Parameters (CPP) for biomanufacturing. Constraint-based systems biology models like Flux Balance Analysis (FBA) are used to estimate metabolic reaction rates, and metabolic rates are used as inputs for multivariate Batch Evolution Models (BEM). Metabolic activity was reproducible among batches and could be monitored to detect a deliberately induced macroscopic process shift (i.e., temperature change). The CMP approach has the potential to enable \"golden batches\" in biomanufacturing processes to be defined from the internal metabolic activity and to aid in detecting process changes that may impact the quality of the product. Overall, the data suggested that monitoring of metabolic activity has promise for biomanufacturing process control.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142495250","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineering GID4 for use as an N-terminal proline binder via directed evolution 通过定向进化将 GID4 改造为 N 端脯氨酸结合剂

IF 3.8 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-25 DOI: 10.1002/bit.28868

Svetlana P. Ikonomova, Bo Yan, Zhiyi Sun, Rachel B. Lyon, Kelly M. Zatopek, John P. Marino, Zvi Kelman

{"title":"Engineering GID4 for use as an N-terminal proline binder via directed evolution","authors":"Svetlana P. Ikonomova, Bo Yan, Zhiyi Sun, Rachel B. Lyon, Kelly M. Zatopek, John P. Marino, Zvi Kelman","doi":"10.1002/bit.28868","DOIUrl":"https://doi.org/10.1002/bit.28868","url":null,"abstract":"Nucleic acid sequencing technologies have gone through extraordinary advancements in the past several decades, significantly increasing throughput while reducing cost. To create similar advancement in proteomics, numerous approaches are being investigated to advance protein sequencing. One of the promising approaches uses N-terminal amino acid binders (NAABs), also referred to as recognizers, that selectively can identify amino acids at the N-terminus of a peptide. However, there are only a few engineered NAABs currently available that bind to specific amino acids and meet the requirements of a biotechnology reagent. Therefore, additional NAABs need to be identified and engineered to enable confident identification and, ultimately, de novo protein sequencing. To fill this gap, a human protein GID4 was engineered to create a NAAB for N-terminal proline (Nt-Pro). While native GID4 binds Nt-Pro, its binding is weak (µmol/L) and greatly influenced by the identity of residues following the Nt-Pro. Through directed evolution, yeast-surface display, and fluorescence-activated cell sorting, we identified sequence variants of GID4 with increased binding response to Nt-Pro. Moreover, variants with an A252V mutation showed a reduced influence from residues in the second and third positions of the target peptide when binding to Nt-Pro. The workflow outlined here is shown to be a viable strategy for engineering NAABs, even when starting from native Nt-binding proteins whose binding is strongly impacted by the identity of residues following Nt-amino acid.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142490160","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Inhibition Control by Continuous Extractive Fermentation Enhances De Novo 2-Phenylethanol Production by Yeast 连续萃取发酵的抑制控制提高了酵母的新 2-苯基乙醇产量

IF 3.8 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-25 DOI: 10.1002/bit.28872

Alessandro Brewster, Arjan Oudshoorn, Marion van Lotringen, Pieter Nelisse, Emily van den Berg, Marijke Luttik, Jean-Marc Daran

{"title":"Inhibition Control by Continuous Extractive Fermentation Enhances De Novo 2-Phenylethanol Production by Yeast","authors":"Alessandro Brewster, Arjan Oudshoorn, Marion van Lotringen, Pieter Nelisse, Emily van den Berg, Marijke Luttik, Jean-Marc Daran","doi":"10.1002/bit.28872","DOIUrl":"https://doi.org/10.1002/bit.28872","url":null,"abstract":"Current microbial cell factory methods for producing chemicals from renewable resources primarily rely on (fed-)batch production systems, leading to the accumulation of the desired product. Industrially relevant chemicals like 2-phenylethanol (2PE), a flavor and fragrance compound, can exhibit toxicity at low concentrations, inhibit the host activity, and negatively impact titer, rate, and yield. Batch liquid-liquid (L-L) In Situ Product Removal (ISPR) was employed to mitigate inhibition effects, but was not found sufficient for industrial-scale application. Here, we demonstrated that continuous selective L-L ISPR provides the solution for maintaining the productivity of de novo produced 2PE at an industrial pilot scale. A unique bioreactor concept called “Fermentation Accelerated by Separation Technology” (FAST) utilizes hydrostatic pressure differences to separate aqueous- and extractant streams within one unit operation, where both production and product extraction take place - allowing for the control of the concentration of the inhibiting compound. Controlled aqueous 2PE levels (0.43 ± 0.02 g kg−1) and extended production times (>100 h) were obtained and co-inhibiting by-product formation was reduced, resulting in a twofold increase of the final product output of batch L-L ISPR approaches. This study establishes that continuous selective L-L ISPR, enabled by FAST, can be applied for more economically viable production of inhibiting products.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142490676","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Real Case Study of 600 m3 Bubble Column Fermentations: Spatially Resolved Simulations Unveil Optimization Potentials for l-Phenylalanine Production With Escherichia coli 600 立方米气泡塔发酵的真实案例研究：空间分辨模拟揭示大肠杆菌生产 l-苯丙氨酸的优化潜力

IF 3.8 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-25 DOI: 10.1002/bit.28869

Yannic Mast, Adel Ghaderi, Ralf Takors

{"title":"Real Case Study of 600 m3 Bubble Column Fermentations: Spatially Resolved Simulations Unveil Optimization Potentials for l-Phenylalanine Production With Escherichia coli","authors":"Yannic Mast, Adel Ghaderi, Ralf Takors","doi":"10.1002/bit.28869","DOIUrl":"https://doi.org/10.1002/bit.28869","url":null,"abstract":"Large-scale fermentations (»100 m³) often encounter concentration gradients which may significantly affect microbial activities and production performance. Reliably investigating such scenarios in silico would allow to optimize bioproduction. But related simulations are very rare in particular for large bubble columns. Here, we pioneer the spatially resolved investigation of a 600 m³ bubble column operating for Escherichia coli based l-phenylalanine fed-batch production. Microbial kinetics are derived from experimental data. Advanced Euler-Lagrange (EL) computational fluid dynamics (CFD) simulations are applied to track individual bubble dynamics that result from a recently developed bubble breakage model. Thereon, the complex nonlinear characteristics of hydrodynamics, mass transfer, and microbial activities are simulated for large scale and compared with real data. As a key characteristic, zones for upriser, downcomer, and circulation cells were identified that dominate mixing and mass transfer. This results in complex gradients of glucose, dissolved oxygen, and microbial rates dividing the bioreactor into sections. Consequently, alternate feed designs are evaluated splitting real feed rates in two feeds at different locations. The opposite reversed installation of feed spots and spargers improved the product synthesis by 6.24% while alternate scenarios increased the growth rate by 11.05%. The results demonstrate how sophisticated, spatially resolved simulations of hydrodynamics, mass transfer, and microbial kinetics help to optimize bioreactors in silico.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142490177","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Improving the catalytic performance of carbonyl reductase based on the functional loops engineering. 基于功能环工程改善羰基还原酶的催化性能。

IF 3.5 2区生物学

Biotechnology and Bioengineering Pub Date : 2024-10-22 DOI: 10.1002/bit.28864

Tao-Shun Zhou, Xiang-Yang Li, Xiao-Jian Zhang, Xue Cai, Zhi-Qiang Liu, Yu-Guo Zheng

{"title":"Improving the catalytic performance of carbonyl reductase based on the functional loops engineering.","authors":"Tao-Shun Zhou, Xiang-Yang Li, Xiao-Jian Zhang, Xue Cai, Zhi-Qiang Liu, Yu-Guo Zheng","doi":"10.1002/bit.28864","DOIUrl":"https://doi.org/10.1002/bit.28864","url":null,"abstract":"Vibegron functions as a potent and selective β3-adrenergic receptor agonist, with its chiral precursor (2S,3R)-aminohydroxy ester (1b) being crucial to its synthesis. In this study, loop engineering was applied to the carbonyl reductase (EaSDR6) from Exiguobacterium algae to achieve an asymmetric reduction of the (rac)-aminoketone ester 1a. The variant M5 (A138L/A190V/S193A/Y201F/N204A) was obtained and demonstrated an 868-fold increase in catalytic efficiency (kcat/Km = 260.3 s-1 mM-1) and a desirable stereoselectivity (>99% enantiomeric excess, e.e.; >99% diastereomeric excess, d.e.) for the target product 1b in contrast to the wild-type EaSDR6 (WT). Structural alignment with WT indicated that loops 137-154 and 182-210 potentially play vital roles in facilitating catalysis and substrate binding. Moreover, molecular dynamics (MD) simulations of WT-1a and M5-1a complex illustrated that M5-1a exhibits a more effective nucleophilic attack distance and more readily adopts a pre-reaction state. The interaction analysis unveiled that M5 enhanced hydrophobic interactions with substrate 1a on cavities A and B while diminishing unfavorable hydrophilic interactions on cavity C. Computational analysis of binding free energies indicated that M5 displayed heightened affinity towards substrate 1a compared to the WT, aligning with its decreased Km value. Under organic-aqueous biphasic conditions, the M5 mutant showed >99% conversion within 12 h with 300 g/L substrate 1a (highest substrate loading as reported). This study enhanced the catalytic performance of carbonyl reductase through functional loops engineering and established a robust framework for the large-scale biosynthesis of the vibegron intermediate.","PeriodicalId":9168,"journal":{"name":"Biotechnology and Bioengineering","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142458170","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0