Kang Min-Kyoung, Zhou Yongjin J, Buijs Nicolaas A, Nielsen Jens
Department of Biology and Biological Engineering, Chalmers University of Technology, Kemivägen 10, 412 96, Gothenburg, Sweden.
Novo Nordisk Foundation Center for Biosustainability, Chalmers University of Technology, 412 96, Gothenburg, Sweden.
Microb Cell Fact. 2017 May 2;16(1):74. doi: 10.1186/s12934-017-0683-z.
Low catalytic activities of pathway enzymes are often a limitation when using microbial based chemical production. Recent studies indicated that the enzyme activity of aldehyde decarbonylase (AD) is a critical bottleneck for alkane biosynthesis in Saccharomyces cerevisiae. We therefore performed functional screening to identify efficient ADs that can improve alkane production by S. cerevisiae.
A comparative study of ADs originated from a plant, insects, and cyanobacteria were conducted in S. cerevisiae. As a result, expression of aldehyde deformylating oxygenases (ADOs), which are cyanobacterial ADs, from Synechococcus elongatus and Crocosphaera watsonii converted fatty aldehydes to corresponding C alkanes and alkenes. The CwADO showed the highest alkane titer (0.13 mg/L/OD) and the lowest fatty alcohol production (0.55 mg/L/OD). However, no measurable alkanes and alkenes were detected in other AD expressed yeast strains. Dynamic expression of SeADO and CwADO under GAL promoters increased alkane production to 0.20 mg/L/OD and no fatty alcohols, with even number chain lengths from C8 to C14, were detected in the cells.
We demonstrated in vivo enzyme activities of ADs by displaying profiles of alkanes and fatty alcohols in S. cerevisiae. Among the AD enzymes evaluated, cyanobacteria ADOs were found to be suitable for alkane biosynthesis in S. cerevisiae. This work will be helpful to decide an AD candidate for alkane biosynthesis in S. cerevisiae and it will provide useful information for further investigation of AD enzymes with improved activities.
在利用微生物进行化学品生产时，途径酶的低催化活性常常是一个限制因素。最近的研究表明，醛脱羰酶（AD）的酶活性是酿酒酵母中烷烃生物合成的关键瓶颈。因此，我们进行了功能筛选，以鉴定能够提高酿酒酵母烷烃产量的高效AD。
对源自植物、昆虫和蓝细菌的AD在酿酒酵母中进行了比较研究。结果，来自聚球藻和沃森球石藻的蓝细菌AD——醛脱甲酰基加氧酶（ADO）的表达将脂肪醛转化为相应的C烷烃和烯烃。CwADO显示出最高的烷烃滴度（0.13 mg/L/OD）和最低的脂肪醇产量（0.55 mg/L/OD）。然而，在其他表达AD的酵母菌株中未检测到可测量的烷烃和烯烃。在GAL启动子下动态表达SeADO和CwADO可将烷烃产量提高到0.20 mg/L/OD，并且在细胞中未检测到碳链长度为C8至C14的偶数链脂肪醇。
我们通过展示酿酒酵母中烷烃和脂肪醇的概况，证明了AD的体内酶活性。在所评估的AD酶中，发现蓝细菌ADO适合酿酒酵母中的烷烃生物合成。这项工作将有助于确定酿酒酵母中烷烃生物合成的AD候选物，并将为进一步研究具有更高活性的AD酶提供有用信息。
