Science Center for Future Foods, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
Key Laboratory of Industrial Biotechnology, Ministry of Education and School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
ACS Synth Biol. 2022 Sep 16;11(9):2979-2988. doi: 10.1021/acssynbio.2c00100. Epub 2022 Aug 17.
Research studies on NAD have proven its crucial role in aging and disease. Nicotinamide mononucleotide (NMN), as the key intermediate of NAD, plays a significant role in supplying and maintaining NAD levels. In the present study, a biocatalytic method for the efficient synthesis of NMN was established. First, was systematically modified to make it more conducive to the biosynthesis and accumulation of NMN. Next, the performance of nicotinamide phosphoribosyltransferase from bacteriophage KVP40 (VpNadV) was determined, which has the best catalytic activity to produce NMN from nicotinamide. The accumulation of extracellular NMN was further increased after the introduction of an NMN transporter. Fine-tuning of gene expression and copy number led to the synthesis of NMN at the yield of 2.6 g/L at the shake flask level. The introduction of a nicotinamide transporter, BcniaP, could not obviously increase the production of NMN at the shake flask level, but it decreased the production of NMN at the bioreactor level. Finally, the titer of NMN reached 16.2 g/L with a conversion ratio of 97.0% from nicotinamide, both of which are highest according to currently available reports. The fed-batch fermentation with direct supplementation of nicotinamide could facilitate the industrial-scale production of NMN compared to that achieved by the whole-cell catalysis process. These results also represent the highest reported yield of NMN synthesized from nicotinamide in .
关于 NAD 的研究已经证明了它在衰老和疾病中的关键作用。烟酰胺单核苷酸（NMN）作为 NAD 的关键中间体，在供应和维持 NAD 水平方面起着重要作用。在本研究中，建立了一种高效合成 NMN 的生物催化方法。首先，系统地修饰 ，使其更有利于 NMN 的生物合成和积累。接下来，测定了来自噬菌体 KVP40（VpNadV）的烟酰胺磷酸核糖基转移酶（Nicotinamide phosphoribosyltransferase）的性能，该酶具有从烟酰胺生产 NMN 的最佳催化活性。引入 NMN 转运蛋白后，进一步增加了细胞外 NMN 的积累。精细调节基因表达和拷贝数可使 NMN 的产量达到摇瓶水平 2.6 g/L。在摇瓶水平上，引入烟酰胺转运蛋白 BcniaP 并不能明显增加 NMN 的产量，但在生物反应器水平上却降低了 NMN 的产量。最后，NMN 的浓度达到 16.2 g/L，转化率为 97.0%，这是目前报道的最高值。与全细胞催化过程相比，直接补加烟酰胺的分批补料发酵更有利于 NMN 的工业化生产。这些结果也代表了目前报道的从烟酰胺合成 NMN 的最高产量。
