A DNA-scaffold platform enhances a multi-enzymatic cycling reaction.
Biotechnol Lett
; 40(4): 667-672, 2018 Apr.
Article
em En
| MEDLINE
| ID: mdl-29359235
OBJECTIVE: We explored the co-localization of multiple enzymes on a DNA backbone via a DNA-binding protein, Gene-A* (A*-tag) to increase the efficiency of cascade enzymatic reactions. RESULTS: Firefly luciferase (FLuc) and pyruvate orthophosphate dikinase (PPDK) were genetically fused with A*-tag and modified with single-stranded (ss) DNA via A*-tag. The components were assembled on ssDNA by hybridization, thereby enhancing the efficiency of the cascading bioluminescent reaction producing light emission from pyrophosphate. The activity of A*-tag in each enzyme was investigated with dye-labeled DNA. Co-localization of the enzymes via hybridization was examined using a gel shift assay. The multi-enzyme complex showed significant improvement in the overall efficiency of the cascading reaction in comparison to a mixture of free enzymes. CONCLUSION: A*-tag is highly convenient for ssDNA modification of versatile enzymes, and it can be used for construction of functional DNA-enzyme complexes.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA de Cadeia Simples
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Piruvato Ortofosfato Diquinase
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Proteínas de Ligação a DNA
Idioma:
En
Ano de publicação:
2018
Tipo de documento:
Article