title: The Development of Leucine Dehydrogenase and Formate Dehydrogenase Bifunctional Enzyme Cascade Improves the Biosynthsis of L-tert-Leucine.
abstract: Leucine dehydrogenase (LDH) and formate dehydrogenase (FDH) were assembled together based on a high-affinity interaction between two different cohesins in a miniscaffoldin and corresponding dockerins in LDH and FDH. The miniscaffoldin with two enzymes was further absorbed by regenerated amorphous cellulose (RAC) to form a bifunctional enzyme complex (miniscaffoldin with LDH and FDH adsorbed by RAC, RSLF) in vitro. The enzymatic characteristics of the bifunctional enzyme complex and free enzymes mixture were systematically compared. The synthesis of L-tert-leucine by the RSLF and free enzyme mixture were compared under different concentrations of enzymes, coenzyme, and substrates. The initial L-tert-leucine production rate by RSLF was enhanced by 2-fold compared with that of the free enzyme mixture. Ninety-one grams per liter of L-tert-leucine with an enantiomeric purity of 99 % e.e. was obtained by RSLF multienzyme catalysis. The results indicated that the bifuntional enzyme complex based on cohesin-dockerin interaction has great potential in the synthesis of L-tert-leucine.
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.
authors.forename: Jixue
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.
authors.forename: Yonghui
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.
authors.forename: Dongfang
authors.initials: D
authors.lastname: Sun
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Jiang
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China. szwang@xmu.edu.cn.|The Key Lab for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen, Fujian, 361005, People's Republic of China. szwang@xmu.edu.cn.
authors.forename: Shizhen
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China. fbs@xmu.edu.cn.|The Key Lab for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen, Fujian, 361005, People's Republic of China. fbs@xmu.edu.cn.
authors.forename: Baishan
authors.initials: B
authors.lastname: Fang
mesh_terms: D000586:Amination; D053898:Biosynthetic Pathways; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005560:Formate Dehydrogenases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007930:Leucine / Q000096:biosynthesis* / Q000737:chemistry; D050803:Leucine Dehydrogenase / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D019289:Pyruvic Acid / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification; D013696:Temperature
chemical_list: D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D009243:NAD; D019289:Pyruvic Acid; D005560:Formate Dehydrogenases; D050803:Leucine Dehydrogenase; D007930:Leucine
keywords: Acceleration factor; Enzyme complex; Formate dehydrogenase; L-tert-leucine; Leucine dehydrogenase; Scaffoldin