(R/S) - 乳酸盐/2-基酸盐在Ralstonia eutropha的区块共聚合物的脱水和生物合成中脱水
Shizuru Ishihara1, Izumi Orita1, Ken'ichiro Matsumoto2
1School of Life Science and Technology, Tokyo Institute of Technology, B-37 4259 Nagatsuta, Midori-ku, Yokohama, 226-8501, Japan.
Applied microbiology and biotechnology
|September 29, 2023
概括
研究人员对Ralstonia eutropha进行了工程设计,以产生新型生物降解的聚酸酸盐 (PHAs) 阻断共聚合物. 这项研究确定了负责 (R/S) -2-基酸盐 (2HB) 敏感性的关键酶,从而实现了增强的PHA合成.
科学领域:
- 生物技术是生物技术.
- 聚合物科学 聚合物科学
- 微生物学 微生物学
背景情况:
- 细菌聚酸酸 (PHAs) 是具有显著潜力的可生物降解聚合物.
- 之前的研究表明,使用大肠杆菌和化学酶 (PhaCAR) 的新型PHA阻断共聚物.
- 自然PHA生产者Ralstonia eutropha被调查了PhaCAR的应用.
研究的目的:
- 在Ralstonia eutropha中应用序列调节PhaCAR酶,用于合成新型PHA阻断共聚合物.
- 为了确定由 (R/S) -2-基酸盐 (2HB) 抑制R. eutropha生长背后的机制.
- 设计R.eutropha以有效生产含有2HB的PHA块共聚合物.
主要方法:
- 在R. eutropha.中表达了仿真PhaCAR酶的表达.
- 关于 (R/S) - 2HB 抑制生长效应的研究和代谢分析.
- 乳酸脱酶同类体的删除分析,以确定关键酶.
- 对R. eutropha的基因工程,包括基因删除和过度表达.
- 在葡萄糖上培育工程菌株,添加 (RS) - 2HB.
- 使用溶剂分化和NMR分析对合成的PHA进行表征.
主要成果:
- (R/S)-2HB对R.eutropha表现出显著的增长抑制作用,与L-valine耗尽有关.
- 细胞染色体依赖的D-乳酸脱酶 (Dld) 和[Fe-S]蛋白依赖的L-乳酸脱酶被确定为分别对 (R) - 2HB和 (S) - 2HB的敏感性负责.
- 工程R.eutropha合成了含有10mol% 2HB的PHA阻断共聚合物,在PhaCAR过度表达时增加到35mol%.
- 由此产生的PHA被证实是块聚合物,表明PhaCAR的序列调节功能与宿主无关.
结论:
- 序列调节的PhaCAR酶可以在Ralstonia eutropha中有效地利用,产生新的PHA阻断共聚合物.
- 了解和克服2HB所造成的代谢限制对于高效的PHA合成至关重要.
- 改造的R. eutropha菌株提供了一个有前途的平台,用于生产定制的可生物降解聚合物,具有可调节的2HB含量.
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