Review
Chemistry, Multidisciplinary
Yajie Wang, Pu Xue, Mingfeng Cao, Tianhao Yu, Stephan T. Lane, Huimin Zhao
Summary: Directed evolution aims to accelerate the natural evolution process of biological molecules and systems through gene diversification and library screening, serving as a powerful tool for engineering improved functions in proteins, metabolic pathways, and whole genomes. Common strategies include gene diversification, screening/selection methods, and continuous evolution, with applications in nucleic acids, proteins, pathways, genetic circuits, viruses, and cells. Challenges and future perspectives in directed evolution are also discussed.
Article
Chemistry, Physical
Zexing Wen, Zhi-Min Zhang, Liang Zhong, Jiaqian Fan, Min Li, Yuanhong Ma, Yang Zhou, Wei Zhang, Bin Guo, Bo Chen, Jian-Bo Wang
Summary: Plant glycosyltransferase MiCGT was engineered by directed evolution to catalyze the glycosylation of flavonoids for pharmaceutical applications, resulting in mutants with enhanced selectivity and activity towards seven different flavonoids. The best quadruple mutant VFAH showed strict 3-O glycosylation selectivity and a 120-fold activity enhancement towards the model substrate quercetin.
Review
Chemistry, Applied
Manfred T. Reetz
Summary: The concept of directed evolution of stereoselective enzymes was born and implemented at the Max-Planck-Institut fur Kohlenforschung in Mulheim, Germany. Andreas Pfaltz witnessed the birth and development of this field during his time at the institute.
ADVANCED SYNTHESIS & CATALYSIS
(2022)
Article
Chemistry, Multidisciplinary
Yang Gu, Brandon Bloomer, Zhennan Liu, Douglas Clark, John F. Hartwig
Summary: Artificial metalloenzymes (ArMs) are a promising class of catalysts created by introducing synthetic cofactors into protein scaffolds for non-natural reactions. Researchers developed a platform to combine mutants of the P450 enzyme CYP119 and the cofactor Ir(Me)MPIX in vivo, leading to the synthesis of chiral amino esters with high TON and good enantioselectivity. The directed evolution of ArMs has been shown to be comparable to that of natural enzymes in vivo.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2022)
Article
Chemistry, Multidisciplinary
Jaeseung Yu, Jinsol Yang, Chaok Seok, Woon Ju Song
Summary: This study demonstrates that protein symmetry can serve as a versatile criterion for identifying promising hotspots for directed evolution of de novo oligomeric enzymes. Randomizing symmetry-related residues located at rotational axes can significantly impact catalytic activities.
Article
Multidisciplinary Sciences
Kazunori Yoshida, Shun Kawai, Masaya Fujitani, Satoshi Koikeda, Ryuji Kato, Tadashi Ema
Summary: By combining loop-walking method and machine learning, a hot-spot loop crucial for protein thermostability was identified, and an ideal mutant was discovered from 214 variants. Though screening the best mutant remains challenging, a computational discrimination model was successfully constructed to predict thermostability, leading to the selection of 20 promising candidates among 7786 combinations for further evaluation. This approach provides a foundation for improvement and optimization in the future.
SCIENTIFIC REPORTS
(2021)
Article
Chemistry, Multidisciplinary
David M. Upp, Rui Huang, Ying Li, Max J. Bultman, Benoit Roux, Jared C. Lewis
Summary: Artificial metalloenzymes (ArMs) were engineered to catalyze diazo cross-coupling resulting in high enantioselectivity alkene reduction to alkane in a one-pot cascade reaction. Directed evolution improved reaction yields and selectivities for various substrates. Molecular dynamics simulations of ArM variants were used to understand the structural role of the cofactor on ArM conformational dynamics, highlighting the ability of ArMs to control both catalyst stereoselectivity and chemoselectivity in complex media.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2021)
Article
Chemistry, Physical
Xiaodong Zhang, Panpan Shen, Jing Zhao, Yueyue Chen, Xian Li, Jian-Wen Huang, Lilan Zhang, Qian Li, Chenghua Gao, Qiong Xing, Chun-Chi Chen, Rey-Ting Guo, Aitao Li
Summary: A highly selective and active β16 steroid hydroxylase CYP109B4 was identified, and the molecular basis for selectivity control was revealed through mutation and computational analysis. This study provides guidance in rational design of more excellent P450 biocatalysts and has potential practical applications.
Article
Chemistry, Physical
Wenlong Zheng, Haoran Yu, Sai Fang, Kaitong Chen, Zhe Wang, Xiuli Cheng, Gang Xu, Lirong Yang, Jianping Wu
Summary: The study utilized a CAST/ISM strategy to accelerate the directed evolution of LTA by focusing on amino acid residues within the substrate pocket. Significant progress was made in the synthesis of products, showing promise for industrial applications.
Article
Chemistry, Physical
Jinlong Li, Sijia Wang, Cui Liu, Yixin Li, Yu Wei, Gang Fu, Pi Liu, Hongwu Ma, Dawei Huang, Jianping Lin, Dawei Zhang
Summary: In this study, a computation-driven strategy for optimizing enzyme structures based on simulation-based iterative saturation mutagenesis (ISM) has been proposed. This strategy aims to stabilize the spatial effects of the transition state (TS) by using a transition state analogue (TSA) as a placeholder. The approach was successfully applied to redesign the structure of chitinase and significantly increase its catalytic activity.
Article
Chemistry, Physical
Sandro Fischer, Thomas R. Ward, Alexandria D. Liang
Summary: Artificial metalloenzymes (ArMs) are created by incorporating synthetic metal catalysts into protein scaffolds, combining the advantages of natural enzymes and synthetic catalysts. The choice of protein scaffold is crucial in tuning the activity of ArMs. The repurposing of the HaloTag protein as an ArM scaffold for metathesis showed promising results in terms of substrate scope and turnover numbers.
Review
Biochemical Research Methods
Zhen Liu, Frances H. Arnold
Summary: Hemoprotein-catalyzed carbene and nitrene transformations are powerful tools for constructing complex molecules, showcasing the emergence and evolution of new protein catalysts. These laboratory-invented enzymes harness the ability of proteins to tame highly reactive carbene and nitrene species with high selectivity. Recent studies have shown that biocatalysts outperform small-molecule catalysts in terms of selectivity and turnover rates.
CURRENT OPINION IN BIOTECHNOLOGY
(2021)
Article
Biochemistry & Molecular Biology
Sarah R. Messenger, Edward M. R. Mcguinniety, Luke J. Stevenson, Jeremy G. Owen, Gregory L. Challis, David F. Ackerley, Mark J. Calcott
Summary: Researchers have developed an evolution-inspired method to generate diverse nonribosomal peptides by amplifying and substituting domains from soil metagenomic DNA.
NATURE CHEMICAL BIOLOGY
(2023)
Article
Chemistry, Physical
Heming Sun, Wenlong Zhu, Qinfei Zhang, Ruonan Zheng, Luo Liu, Hui Cao
Summary: Researchers modified and designed polyphosphate-dependent glucomannokinase (PPGMK) to utilize inorganic polyphosphate for synthesis, and found an optimal combination mutant with improved thermal stability, longer half-life, and increased enzyme activity.
Article
Chemistry, Multidisciplinary
Claire G. Page, Simon J. Cooper, Jacob S. DeHovitz, Daniel G. Oblinsky, Kyle F. Biegasiewicz, Alyssa H. Antropow, Kurt W. Armbrust, J. Michael Ellis, Lawrence G. Hamann, Evan J. Horn, Kevin M. Oberg, Gregory D. Scholes, Todd K. Hyster
Summary: This study presents a photoenzymatic intermolecular hydroalkylation of olefins catalyzed by flavin-dependent 'ene'-reductases. A unique mechanism involving a charge-transfer complex formed between an alkene, alpha-chloroamide, and flavin hydroquinone ensures radical formation only occurs when both substrates are present within the protein active site. This work demonstrates the potential for new biocatalytic transformations enabled by photoenzymatic catalysis through previously unknown electron transfer mechanisms.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2021)
