合成支架在代谢工程中的研究进展
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摘要
代谢工程作为通过引入外源合成途径或改造优化代谢网络,进行高附加值的天然代谢产物生物合成的技术,已经得到广泛应用。但随着目标合成产物的结构日渐复杂,构建多基因的从头合成途径造成宿主生物代谢失衡与中间产物对宿主细胞产生毒害作用等一系列问题发生的可能性也随之增加。为解决这些问题合成支架策略应运而生,合成支架将途径酶共定位以提高局部酶和代谢物的浓度,来增强代谢通量并限制中间产物与宿主细胞环境间的相互作用,成为生物催化和合成生物学研究的热点之一。尽管由核酸、蛋白质构成的合成支架策略已经应用于多种代谢物的异源合成,并取得了不同程度的成功,但合成支架的精确组装仍然是一项艰巨的任务。文中详细介绍了合成支架技术的研究现状,详细阐述了合成支架技术的原理和实例,并初步探讨了其应用前景。
Metabolic engineering is a powerful tool to increase many valuable metabolites through enhancing endogenous pathways or introducing exogenous pathways from other organisms. As the complexity of the targeted structure increases,many problems arise when the host suffers from flux imbalance and some toxic effects. An emerging approach to solve these problems is the use of synthetic scaffolds to co-localize key enzymes and metabolites of the synthetic pathways, enhance the metabolic flux and limit the interaction between intermediate products in the host cell. Although many scaffolds made of proteins and nucleic acids have been explored and applied to a variety of research to the heterogeneous synthesis of multiple metabolites, success is rather limited. The precise assembly of synthetic scaffolds remains a difficult task. In this review, we summarized the application of synthetic scaffolds in metabolic engineering, and outlined the main principle of scaffold designs, then highlighted the current challenges in their application.
Metabolic engineering is a powerful tool to increase many valuable metabolites through enhancing endogenous pathways or introducing exogenous pathways from other organisms. As the complexity of the targeted structure increases,many problems arise when the host suffers from flux imbalance and some toxic effects. An emerging approach to solve these problems is the use of synthetic scaffolds to co-localize key enzymes and metabolites of the synthetic pathways, enhance the metabolic flux and limit the interaction between intermediate products in the host cell. Although many scaffolds made of proteins and nucleic acids have been explored and applied to a variety of research to the heterogeneous synthesis of multiple metabolites, success is rather limited. The precise assembly of synthetic scaffolds remains a difficult task. In this review, we summarized the application of synthetic scaffolds in metabolic engineering, and outlined the main principle of scaffold designs, then highlighted the current challenges in their application.
引文
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[34]Hirakawa H,Nagamune T.Molecular assembly of P450with ferredoxin and ferredoxin reductase by fusion to PCNA.Chembiochem,2010,11(11):1517-1520.
[35]Hirakawa H,Kakitani A,Nagamune T.Introduction of selective intersubunit disulfide bonds into self-assembly protein scaffold to enhance an artificial multienzyme complex’s activity.Biotechnol Bioeng,2013,110(7):1858-1864.
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[37]Moon TS,Dueber JE,Shiue E,et al.Use of modular,synthetic scaffolds for improved production of glucaric acid in engineered E.coli.Metab Eng,2010,12(3):298-305.
[38]Agapakis CM,Ducat DC,Boyle PM,et al.Insulation of a synthetic hydrogen metabolism circuit in bacteria.J Biol Eng,2010,4:3.
[39]Baek JM,Mazumdar S,Lee SW,et al.Butyrate production in engineered Escherichia coli with synthetic scaffolds.Biotechnol Bioeng,2013,110(10):2790-2794.
[40]Wang YC,Yu O.Synthetic scaffolds increased resveratrol biosynthesis in engineered yeast cells.JBiotechnol,2012,157(1):258-260.
[41]Gao X,Yang S,Zhao CC,et al.Artificial multienzyme supramolecular device:highly ordered self-assembly of oligomeric enzymes in vitro and in vivo.Angew Chem Int Ed Engl,2015,53(51):14027-14030.
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[43]Yang ZW,Gao X,Xie H,et al.Enhanced itaconic acid production by self-assembly of two biosynthetic enzymes in Escherichia coli.Biotechnol Bioeng,2016,114(2):457-462.
[44]Somasundaram S,Eom GT,Hong SH.Efficient malic acid production in Escherichia coli using a synthetic scaffold protein complex.Appl Biochem Biotechnol,2017,184(4):1308-1318.
[45]Zhang L,Lu Y,Zhong YQ.Affibody molecules:a new class of ligands with high affinity.J Int Pharm Res,2012,39(2):127-131(in Chinese).张磊,鲁莹,钟延强.Affibody分子:一类具有高度亲和力的新配体.国际药学研究杂志,2012,39(2):127-131.
[46]Tippmann S,Anfelt J,David F,et al.Affibody Scaffolds improve sesquiterpene production in Saccharomyces cerevisiae.ACS Synth Biol,2016,6(1):19-28.
[47]Shetty RP,Endy D,Knight Jr TF.Engineering biobrick vectors from biobrick parts.J Biol Eng,2008,2:5.
[48]Jaeger L,Chworos A.The architectonics of programmable RNA and DNA nanostructures.Curr Opin Struc Biol,2006,16(4):531-543.
[49]Rothemund PW.Folding DNA to create nanoscale shapes and patterns.Nature,2006,440(7082):297-302.
[50]Douglas SM,Dietz H,Liedl T,et al.Self-assembly of DNA into nanoscale three-dimensional shapes.Nature,2010,459(7245):414-418.
[51]Han DR,Pal S,Nangreave J,et al.DNA origami with complex curvatures in three-dimensional space.Science,2011,332(6027):342-346.
[52]Zhang F,Jiang SX,Wu SY,et al.Complex wireframe DNA origami nanostructures with multi-arm junction vertices.Nat Nanotechnol,2015,10(9):779-784.
[53]Zuker M.Mfold web server for nucleic acid folding and hybridization prediction.Nucleic Acids Res,2003,31(13):3406-3415.
[54]Zadeh JN,Steenberg CD,Bois JS,et al.NUPACK:Analysis and design of nucleic acid systems.JComput Chem,2011,32(1):170-173.
[55]Andronescu M,Aguirre-Hernandez R,Condon A,et al.RNAsoft:a suite of RNA secondary structure prediction and design software tools.Nucleic Acids Res,2003,31(13):3416-3422.
[56]Douglas SM,Marblestone AH,Teerapittayanon S,et al.Rapid prototyping of 3D DNA-origami shapes with caDNAno.Nucleic Acids Res,2009,37(15):5001-5006.
[57]Castro CE,Kilchherr F,Kim DN,et al.A primer to scaffolded DNA origami.Nat Methods,2011,8(3):221-229.
[58]Williams S,Lund K,Lin CX,et al.Tiamat:a three-dimensional editing tool for complex DNAstructures//Goel A,Simmel FC,Sosík P,eds.DNAComputing.New York:Springer Berlin Heidelberg,2008,5347:90-101.
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