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Biocatalysis pp 167-176 | Cite as

Screening, Optimization and Assembly of Key Pathway Enzymes in Metabolic Engineering

  • Yanfeng Liu
  • Long LiuEmail author
Chapter

Abstract

Metabolic engineering is an enabling technology for producing chemicals, pharmaceuticals, and fuels in a green and sustainable manner. Enzymes are key catalysts for metabolic reactions for the synthesis of target product. In this chapter, we initially discuss enzyme research in metabolic engineering, fueled by screening the enzymes of key biochemical pathways from different organisms for an enhanced production. Next, the optimization of key pathway enzymes by feedback inhibition removal, catalytic efficiency improvement, and substrate specificity alteration are discussed. Finally, assembling of the key pathway enzymes for balancing and strengthening synthetic pathways is discussed, including fusion expression of key enzymes, synthetic scaffold-guided enzyme co-localization, and compartmentalization engineering. The systematic summary and discussion of screening, optimization and assembling of key pathway enzymes in metabolic engineering may facilitate metabolic engineers to further combine protein engineering with metabolic engineering for eliminating rate-limiting steps for improved production.

Keywords

Metabolic engineering Green chemistry Biochemical production Pathway enzymes 

References

  1. Alper H, Fischer C, Nevoigt E, Stephanopoulos G (2006) Tuning genetic control through promoter engineering (vol 102, pg 12678, 2005). Proc Natl Acad Sci U S A 103(8):3006–3006CrossRefGoogle Scholar
  2. Arifuzzaman M, Maeda M, Itoh A, Nishikata K, Takita C, Saito R, Ara T, Nakahigashi K, Huang HC, Hirai A (2006) Large-scale identification of protein–protein interaction of Escherichia coli K-12. Genome Res 16(5):686–691PubMedPubMedCentralCrossRefGoogle Scholar
  3. Avalos JL, Fink GR, Stephanopoulos G (2013) Compartmentalization of metabolic pathways in yeast mitochondria improves the production of branched-chain alcohols. Nat Biotechnol 31(4):335–341PubMedPubMedCentralCrossRefGoogle Scholar
  4. Becker J, Wittmann C (2015) Advanced biotechnology: metabolically engineered cells for the bio-based production of chemicals and fuels, materials, and health-care products. Angew Chem Int Ed 54(11):3328–3350CrossRefGoogle Scholar
  5. Castellana M, Wilson MZ, Xu Y, Joshi P, Cristea IM, Rabinowitz JD, Gitai Z, Wingreen NS (2014) Enzyme clustering accelerates processing of intermediates through metabolic channeling. Nat Biotechnol 32(10):1011–1018PubMedPubMedCentralCrossRefGoogle Scholar
  6. Chen L, Chen Z, Zheng P, Sun J, Zeng AP (2012) Study and reengineering of the binding sites and allosteric regulation of biosynthetic threonine deaminase by isoleucine and valine in Escherichia coli. Appl Microbiol Biotechnol 97(7):2939–2949PubMedCrossRefGoogle Scholar
  7. Chen Z, Bommareddy RR, Frank D, Rappert S, Zeng A-P (2014a) Deregulation of feedback inhibition of phosphoenolpyruvate carboxylase for improved lysine production in Corynebacterium glutamicum. Appl Environ Microbiol 80(4):1388–1393PubMedPubMedCentralCrossRefGoogle Scholar
  8. Chen Z, Rappert S, Zeng A-P (2014b) Rational design of allosteric regulation of homoserine dehydrogenase by a non-natural inhibitor L-lysine. ACS Synth Biol 4(2):126–131PubMedCrossRefGoogle Scholar
  9. Chen X, Zhou J, Zhang L, Pu Z, Liu L, Shen W, Fan Y (2018) Development of an Escherichia coli-based biocatalytic system for the efficient synthesis of N-acetyl-D-neuraminic acid. Metab Eng 47:374–382PubMedCrossRefGoogle Scholar
  10. Conrado RJ, Wu GC, Boock JT, Xu H, Chen SY, Lebar T, Turnšek J, Tomšič N, Avbelj M, Koprivnjak T (2012) DNA-guided assembly of biosynthetic pathways promotes improved catalytic efficiency. Nucleic Acids Res 40(4):1879–1889PubMedCrossRefGoogle Scholar
  11. Delebecque CJ, Silver PA, Lindner AB (2012) Designing and using RNA scaffolds to assemble proteins in vivo. Nat Protoc 7(10):1797–1807PubMedCrossRefGoogle Scholar
  12. DeLoache WC, Russ ZN, Narcross L, Gonzales AM, Martin VJ, Dueber JE (2015) An enzyme-coupled biosensor enables (S)-reticuline production in yeast from glucose. Nat Chem Biol 11(7):465–471CrossRefGoogle Scholar
  13. Deng MD, Severson DK, Grund AD, Wassink SL, Burlingame RP, Berry A, Running JA, Kunesh CA, Song L, Jerrell TA (2005) Metabolic engineering of Escherichia coli for industrial production of glucosamine and N-acetylglucosamine. Metab Eng 7(3):201–214PubMedCrossRefGoogle Scholar
  14. Dueber JE, Wu GC, Malmirchegini GR, Moon TS, Petzold CJ, Ullal AV, Prather KLJ, Keasling JD (2009) Synthetic protein scaffolds provide modular control over metabolic flux. Nat Biotechnol 27(8):753–759PubMedPubMedCentralCrossRefGoogle Scholar
  15. Galanie S, Thodey K, Trenchard IJ, Filsinger Interrante M, Smolke CD (2015) Complete biosynthesis of opioids in yeast. Science 349(6252):1095–1100PubMedPubMedCentralCrossRefGoogle Scholar
  16. Gerosa L, Sauer U (2011) Regulation and control of metabolic fluxes in microbes. Curr Opin Biotechnol 22(4):566–575PubMedCrossRefGoogle Scholar
  17. Hu P, Janga SC, Babu M, Díazmejía JJ, Butland G, Yang W, Pogoutse O, Guo X, Phanse S, Wong P (2009) Global functional atlas of Escherichia coli encompassing previously uncharacterized proteins. PLoS Biol 7(4):e96PubMedCrossRefGoogle Scholar
  18. Lee JW, Na D, Park JM, Lee J, Choi S, Lee SY (2012) Systems metabolic engineering of microorganisms for natural and non-natural chemicals. Nat Chem Biol 8(6):536–546PubMedCrossRefGoogle Scholar
  19. Lee JH, Jung S-C, Kang KH, Song J-J, Kim SC (2013) Improved production of l-threonine in Escherichia coli by use of a DNA scaffold system. Appl Environ Microbiol 79(3):774–782PubMedPubMedCentralCrossRefGoogle Scholar
  20. Lee MJ, Mantell J, Hodgson L, Alibhai D, Fletcher JM, Brown IR, Frank S, Xue W-F, Verkade P, Woolfson DN, Warren MJ (2018) Engineered synthetic scaffolds for organizing proteins within the bacterial cytoplasm. Nat Chem Biol 14(2):142–147PubMedCrossRefGoogle Scholar
  21. Leonard E, Ajikumar PK, Thayer K, Xiao W-H, Mo JD, Tidor B, Stephanopoulos G, Prather KL (2010) Combining metabolic and protein engineering of a terpenoid biosynthetic pathway for overproduction and selectivity control. Proc Natl Acad Sci U S A 107(31):13654–13659PubMedPubMedCentralCrossRefGoogle Scholar
  22. Liu Y, Zhu Y, Ma W, Shin H-D, Li J, Liu L, Du G, Chen J (2014) Spatial modulation of key pathway enzymes by DNA-guided scaffold system and respiration chain engineering for improved N-acetylglucosamine production by Bacillus subtilis. Metab Eng 24:61–69PubMedCrossRefGoogle Scholar
  23. McNerney MP, Watstein DM, Styczynski MP (2015) Precision metabolic engineering: the design of responsive, selective, and controllable metabolic systems. Metab Eng 31:123–131PubMedPubMedCentralCrossRefGoogle Scholar
  24. Nielsen J, Keasling JD (2016) Engineering cellular metabolism. Cell 164(6):1185–1197PubMedCrossRefGoogle Scholar
  25. Nowroozi FF, Baidoo EE, Ermakov S, Redding-Johanson AM, Batth TS, Petzold CJ, Keasling JD (2013) Metabolic pathway optimization using ribosome binding site variants and combinatorial gene assembly. Appl Microbiol Biotechnol 98(4):1567–1581PubMedCrossRefGoogle Scholar
  26. Paddon C, Westfall P, Pitera D, Benjamin K, Fisher K, McPhee D, Leavell M, Tai A, Main A, Eng D (2013) High-level semi-synthetic production of the potent antimalarial artemisinin. Nature 496(7446):528–532PubMedCrossRefGoogle Scholar
  27. Philp JC, Ritchie RJ, Allan JE (2013) Biobased chemicals: the convergence of green chemistry with industrial biotechnology. Trends Biotechnol 31(4):219–222PubMedCrossRefGoogle Scholar
  28. Rajagopala SV, Sikorski P, Kumar A, Mosca R, Vlasblom J, Arnold R, Francakoh J, Pakala SB, Phanse S, Ceol A (2014) The binary protein-protein interaction landscape of Escherichia coli. Nat Biotechnol 32(3):285–290PubMedPubMedCentralCrossRefGoogle Scholar
  29. Rodriguez GM, Tashiro Y, Atsumi S (2014) Expanding ester biosynthesis in Escherichia coli. Nat Chem Biol 10(4):259–265PubMedPubMedCentralCrossRefGoogle Scholar
  30. Thodey K, Galanie S, Smolke CD (2014) A microbial biomanufacturing platform for natural and semisynthetic opioids. Nat Chem Biol 10(10):837–844PubMedPubMedCentralCrossRefGoogle Scholar
  31. Woolston BM, Edgar S, Stephanopoulos G (2013) Metabolic engineering: past and future. Annu Rev Chem Biomol Eng 4:259–288PubMedCrossRefGoogle Scholar
  32. Zhang Y-HP (2011) Substrate channeling and enzyme complexes for biotechnological applications. Biotechnol Adv 29(6):715–725PubMedCrossRefGoogle Scholar
  33. Zhang Y, Li SZ, Li J, Pan X, Cahoon RE, Jaworski JG, Wang X, Jez JM, Chen F, Yu O (2006) Using unnatural protein fusions to engineer resveratrol biosynthesis in yeast and mammalian cells. J Am Chem Soc 128(40):13030–13031PubMedCrossRefGoogle Scholar
  34. Zhang X, Liu Y, Liu L, Wang M, Li J, Du G, Chen J (2018) Modular pathway engineering of key carbon-precursor supply-pathways for improved N-acetylneuraminic acid production in Bacillus subtilis. Biotechnol Bioeng 115(9):2217–2231PubMedCrossRefGoogle Scholar
  35. Zhou YJ, Gao W, Rong Q, Jin G, Chu H, Liu W, Yang W, Zhu Z, Li G, Zhu G, Huang L, Zhao ZK (2012) Modular pathway engineering of diterpenoid synthases and the mevalonic acid pathway for miltiradiene production. J Am Chem Soc 134(6):3234–3241PubMedCrossRefGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of EducationJiangnan UniversityWuxiChina
  2. 2.Key Laboratory of Industrial Biotechnology, Ministry of EducationJiangnan UniversityWuxiChina

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