Department of Microbiology and Genetics, Berlin University of Technology, Seestr. 13, D-13353 Berlin, Germany.
Metab Eng. 2009 Nov;11(6):335-46. doi: 10.1016/j.ymben.2009.07.005. Epub 2009 Jul 24.
Dihydroxyacetone (DHA) has numerous industrial applications. In this work, we pursue the idea to produce DHA from sugars in the yeast Saccharomyces cerevisiae, via glycerol as an intermediate. Firstly, three glycerol dehydrogenase (GDH) genes from different microbial sources were expressed in yeast. Among them, the NAD(+)-dependent GDH of Hansenula polymorpha showed the highest glycerol-oxidizing activity. DHA concentration in shake-flask experiments was roughly 100mg/lDHA from 20g/l glucose, i.e. five times the wild-type level. This level was achieved only when cultures were subjected to osmotic stress, known to enhance glycerol production and accumulation in S. cerevisiae. Secondly, DHA kinase activity was abolished to prevent conversion of DHA to dihydroxyacetone phosphate (DHAP). The dak1Deltadak2Delta double-deletion mutant overexpressing H. polymorpha gdh produced 700mg/l DHA under the same conditions. Although current DHA yield and titer still need to be optimized, our approach provides the proof of concept for producing DHA from sugars in yeast.
二羟丙酮（DHA）有许多工业用途。在这项工作中，我们设想通过甘油作为中间体，从酵母酿酒酵母中的糖中生产 DHA。首先，从不同微生物来源表达了三种甘油脱氢酶（GDH）基因。其中，粘红酵母的 NAD(+)依赖性 GDH 表现出最高的甘油氧化活性。在摇瓶实验中，从 20g/l 葡萄糖中获得的 DHA 浓度约为 100mg/lDHA，即比野生型水平高五倍。只有当培养物受到渗透胁迫时，才能达到这种水平，已知渗透胁迫会促进酿酒酵母中甘油的产生和积累。其次，为了防止 DHA 转化为二羟丙酮磷酸（DHAP），消除了 DHA 激酶活性。在相同条件下，过表达粘红酵母 gdh 的 dak1Deltadak2Delta 双缺失突变体产生了 700mg/l DHA。尽管目前的 DHA 产量和浓度仍需优化，但我们的方法为从酵母中的糖中生产 DHA 提供了概念验证。
