title: Enhancement of the activity of enzyme immobilized on polydopamine-coated iron oxide nanoparticles by rational orientation of formate dehydrogenase.
abstract: Immobilization of enzymes onto nanoparticles and retention of their structure and activity, which may be related to the orientation of enzymes on nanoparticles, remain a challenge. Here, we developed a novel enzyme-orientation strategy to enhance the activity of formate dehydrogenase immobilized on polydopamine-coated iron oxide nanoparticles via site-directed mutation. Seven mutants were constructed based on homology modeling of formate dehydrogenase and immobilized on polydopamine-coated iron oxide nanoparticles to investigate the influence of these mutations on immobilization. The immobilized mutant C242A/C275V/C363V/K389C demonstrated the highest immobilization yield and retained 90% of its initial activity, which was about 3-fold higher than that of wild-type formate dehydrogenase. Moreover, co-immobilization of formate dehydrogenase and leucine dehydrogenase was performed for the synthesis of l-tert-leucine. The catalytic efficiency of the co-immobilized mutant C242A/C275V/C363V/K389C and leucine dehydrogenase increased by more than 4-fold compared to that of co-immobilized wild-type formate dehydrogenase and leucine dehydrogenase.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing Technology, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Gao
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing Technology, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Kefeng
authors.initials: K
authors.lastname: Ni
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing Technology, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Chengcheng
authors.initials: C
authors.lastname: Zhao
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing Technology, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China. Electronic address: yhren@ecust.edu.cn.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Ren
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Shanghai Collaborative Innovation Center for Biomanufacturing Technology, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China. Electronic address: dzhwei@ecust.edu.cn.
authors.forename: Dongzhi
authors.initials: D
authors.lastname: Wei
mesh_terms: D004800:Enzymes, Immobilized / Q000235:genetics / Q000378:metabolism*; D005290:Ferric Compounds / Q000737:chemistry*; D005560:Formate Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D007211:Indoles / Q000737:chemistry*; D053768:Metal Nanoparticles; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D016297:Mutagenesis, Site-Directed; D011108:Polymers / Q000737:chemistry*
chemical_list: D004800:Enzymes, Immobilized; D005290:Ferric Compounds; D007211:Indoles; D011108:Polymers; C568283:polydopamine; C000499:ferric oxide; D005560:Formate Dehydrogenases
keywords: Directional immobilization; Formate dehydrogenase; Iron oxide nanoparticles; Polydopamine; Site-directed mutagenesis