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.

IF 3.5 2区 生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY 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
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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.

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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。
黄素依赖性卤化酶(FDH)因其能够以精确的区域选择性生产卤代芳香族化合物而备受制药和化工行业的青睐。本研究设计了一种多酶自组装策略,为 FDH 构建了一个有效可靠的体外辅酶循环系统。最初,研究人员开发了由葡萄糖脱氢酶(GDH)、黄素还原酶(FR)和FDH组成的三酶自组装纳米簇(TESNCs)。在将 L-Trp 转化为 6-Cl-L-Trp 的过程中,与游离三酶混合物相比,TESNCs 表现出更高的热稳定性和转化效率,使产量增加了 2.1 倍。随后,建立了 GDH、FR 和 FDH 的有序共固定,进一步提高了 FDH 辅酶循环系统的稳定性和催化效率。与游离 TESNCs 相比,固定化 TESNCs 在 5 mL 反应体系中的催化效率提高了 4.2 倍。这项研究为开发稳健高效的 FDH 辅酶循环系统提供了有效策略。
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来源期刊
Biotechnology and Bioengineering
Biotechnology and Bioengineering 工程技术-生物工程与应用微生物
CiteScore
7.90
自引率
5.30%
发文量
280
审稿时长
2.1 months
期刊介绍: Biotechnology & Bioengineering publishes Perspectives, Articles, Reviews, Mini-Reviews, and Communications to the Editor that embrace all aspects of biotechnology. These include: -Enzyme systems and their applications, including enzyme reactors, purification, and applied aspects of protein engineering -Animal-cell biotechnology, including media development -Applied aspects of cellular physiology, metabolism, and energetics -Biocatalysis and applied enzymology, including enzyme reactors, protein engineering, and nanobiotechnology -Biothermodynamics -Biofuels, including biomass and renewable resource engineering -Biomaterials, including delivery systems and materials for tissue engineering -Bioprocess engineering, including kinetics and modeling of biological systems, transport phenomena in bioreactors, bioreactor design, monitoring, and control -Biosensors and instrumentation -Computational and systems biology, including bioinformatics and genomic/proteomic studies -Environmental biotechnology, including biofilms, algal systems, and bioremediation -Metabolic and cellular engineering -Plant-cell biotechnology -Spectroscopic and other analytical techniques for biotechnological applications -Synthetic biology -Tissue engineering, stem-cell bioengineering, regenerative medicine, gene therapy and delivery systems The editors will consider papers for publication based on novelty, their immediate or future impact on biotechnological processes, and their contribution to the advancement of biochemical engineering science. Submission of papers dealing with routine aspects of bioprocessing, description of established equipment, and routine applications of established methodologies (e.g., control strategies, modeling, experimental methods) is discouraged. Theoretical papers will be judged based on the novelty of the approach and their potential impact, or on their novel capability to predict and elucidate experimental observations.
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