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High level expression of nicotinamide nucleoside kinase from Saccharomyces cerevisiae and its purification and immobilization by one-step method

Nicotinamide riboside kinase (NRK) plays an important role in the synthesis of β -nicotinamide nucleotide (NMN). NMN is a key intermediate of NAD+ synthesis, and it actually contribute to the well-being of our health. In this study, gene mining technology was used to clone nicotinamide nucleoside kinase gene fragments from S. cerevisiae, and the ScNRK1 was achieved a high level of soluble expression in E. coli BL21. Then, the reScNRK1 was immobilized by metal affinity label to optimize the enzyme performance. The results showed that the enzyme activity in the fermentation broth was 14.75 IU/mL, and the specific enzyme activity after purification was 2252.59 IU/mg. After immobilization, the optimum temperature of the immobilized enzyme was increased by 10°C compared with the free enzyme, and the temperature stability was improved with little change in pH. Moreover, the activity of the immobilized enzyme remained above 80% after four cycles of immobilized reScNRK1, which makes the enzyme more advantageous in the enzymatic synthesis of NMN.

 

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This study describes the cloning and optimization of the nicotinamide nucleoside kinase gene from S. cerevisiae, ScNRK1, to improve the enzyme performance for the synthesis of β-nicotinamide nucleotide (NMN). The researchers achieved a high level of soluble expression of ScNRK1 in E. coli BL21 and then immobilized the reScNRK1 using metal affinity labeling to optimize its performance.

The study found that the enzyme activity in the fermentation broth was 14.75 IU/mL, and the specific enzyme activity after purification was 2252.59 IU/mg. After immobilization, the optimum temperature of the enzyme was increased by 10°C compared to the free enzyme, and its temperature stability was improved with little change in pH. Additionally, the immobilized enzyme retained over 80% of its activity after four cycles, making it more advantageous for the enzymatic synthesis of NMN.

Overall, this study provides valuable information on the optimization of ScNRK1 for the synthesis of NMN, which has important implications for human health. NMN is a key intermediate in the synthesis of NAD+ and has been shown to have anti-aging and anti-inflammatory effects in animal studies. The use of immobilized enzymes could also have practical applications in the production of NMN for commercial use.

Related Products

Cat.No. Product Name Information
S5259 β-Nicotinamide Mononucleotide Nicotinamide mononucleotide (β-NM, NMN), one of the key precursors of NAD+ and products of the NAMPT reaction , is converted to NAD+ by nicotinamide mononucleotide adenylyltransferase.

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