Department of Biochemistry and Molecular Biology, University of Bucharest, Bucharest, 050095, Romania.
Independent Research Association, Bucharest, 012416, Romania.
Sci Rep. 2018 Aug 16;8(1):12278. doi: 10.1038/s41598-018-30792-0.
Diabetes is a chronic and progressive disease with continuously increasing prevalence, rising financial pressure on the worldwide healthcare systems. Recently, the insulin resistance, hallmark of type 2 diabetes, was cured in mice treated with NAD precursor β-nicotinamide mononucleotide (NMN), no toxic effects being reported. However, NMN has a high price tag, more cost effective production methods are needed. This study proposes a biotechnological NMN production method in Escherichia coli. We show that bicistronic expression of recombinant nicotinamide phosphoribosyl transferase (Nampt) and phosphoribosyl pyrophosphate (PRPP) synthetase in the presence of nicotinamide (NAM) and lactose may be a successful strategy for cost effective NMN production. Protein expression vectors carrying NAMPT gene from Haemophilus ducreyi and PRPP synthetase from Bacillus amyloliquefaciens with L135I mutation were transformed in Escherichia coli BL21(DE3)pLysS. NMN production reached a maximum of 15.42 mg per L of bacterial culture (or 17.26 mg per gram of protein) in these cells grown in PYA8 medium supplemented with 0.1% NAM and 1% lactose.
糖尿病是一种慢性、渐进性疾病，其患病率不断上升，给全球医疗保健系统带来了不断增加的财政压力。最近，用烟酰胺单核苷酸（NMN）预处理的胰岛素抵抗（2 型糖尿病的标志）在小鼠中得到了治愈，而且没有报道有毒副作用。然而，NMN 的价格很高，需要更具成本效益的生产方法。本研究提出了一种在大肠杆菌中生产 NMN 的生物技术方法。我们表明，在烟酰胺（NAM）和乳糖存在的情况下，双顺反子表达重组烟酰胺磷酸核糖转移酶（Nampt）和磷酸核糖焦磷酸（PRPP）合成酶可能是一种具有成本效益的 NMN 生产的成功策略。携带有 Haemophilus ducreyi 的 Nampt 基因和 Bacillus amyloliquefaciens 的 L135I 突变 PRPP 合成酶的蛋白表达载体被转化到大肠杆菌 BL21(DE3)pLysS 中。在 PYA8 培养基中培养这些细胞，补充 0.1%NAM 和 1%乳糖，NMN 的产量最高达到每升细菌培养物 15.42 毫克（或每克蛋白 17.26 毫克）。
