Journal
JOURNAL OF BIOTECHNOLOGY
Volume 245, Issue -, Pages 14-20Publisher
ELSEVIER
DOI: 10.1016/j.jbiotec.2017.01.010
Keywords
Transaminase; D-amino acid oxidase; Elastin-like polypeptide; MnO2 nanorods; Immobilization
Categories
Funding
- National Science Foundation of China [21376021, 21576018]
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R-omega-transaminases transfer an amino group from an amino donor (e.g. (R)-1-phenylethylamine) onto an amino acceptor (e.g. pyruvate), resulting a co-product (e.g. D-alanine). This work intends to immobilize R-omega-Transaminase on MnO2 nanorods to achieve multienzyme catalysis. R-omega-Transaminase (RTA) and D-amino acid oxidase (DAAO) have been fused to an elastin-like polypeptide (ELP) separately through genetic engineering of the enzymes. ELP-RTA and ELP-DAAO have been separately immobilized on polydopamine-coated MnO2 nanorods. When the two immobilized enzymes were used together in one pot, the transformation of (R)-1-phenylethylamine was catalyzed by the immobilized ELP-RTA, and the co-product D-alanine was converted back to pyruvate under the catalysis of the immobilized ELP-DAAO, achieving the recycling of pyruvate in situ. Thus pyruvate was maintained at a low concentration in order to reduce its negative effect. On the other hand, the generated H2O2 of ELP-DAAO was decomposed by the MnO2 nanorods, and the evolved oxygen oxidized the reduced cofactors of ELP-DAAO. Forming the circles of hydrogen peroxide -> oxygen -> hydrogen peroxide accelerated the deamination reaction. The highly efficient conversion of the co-product D-alanine back to pyruvate accelerated the forming of the pyruvate -> D-alanine -> pyruvate cycle between the two immobilized enzymes. The coordination of the pyruvate -> D-alanine -> pyruvate and hydrogen peroxide oxygen -4 hydrogen peroxide cycles accelerated the transformation of (R)-1-phenylethylamine. As a result, As a result, the immobilized enzymes achieved a conversion of 98 +/- 1.8% in comparison to 69.6 +/- 1.2% by free enzymes. (C) 2017 Elsevier B.V. All rights reserved.
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