奇迹束丝放线菌合成安丝菌素过程中支链氨基酸的添加效果分析

Effects Analysis of Adding Branched Amino Acids on the Biosynthesis of Ansamitocins by Actinosynnema mirum

安丝菌素是一类重要的聚酮抗生素,由AP-1,AP-2,AP-3,AP-3′和AP-4 5个组分组成,其中AP-3的活性最强,通常作为发酵的目标产物。奇迹束丝放线菌合成安丝菌素的摇瓶实验发现,添加支链氨基酸(BCAAs)可明显影响安丝菌素的产量以及组分。为了进一步验证支链氨基酸对安丝菌素生物合成的作用,选用合成培养基并在不同时段添加支链氨基酸进行发酵实验,实验结果显示,发酵48h后加入0.5g/L缬氨酸(Val)能使AP-3增产1倍左右;而在同一时刻添加等量的亮氨酸(Leu)和异亮氨酸(Ile)对AP-3的抑制作用较明显,产量分别下降了58.8%和83.3%,且添加Leu和Ile分别使得AP-4与AP-2组分增加。此外,添加Ile可使琥珀酸与乙酸的积累量都明显上升,而添加Val使得细胞分泌琥珀酸和乙酸明显变少,这意味着奇迹束丝放线菌代谢支链氨基酸不仅影响对聚酮途径以及侧链合成,对细胞的代谢状态也有一定影响。

Ansamitocins are a group of macrolide antibiotics, which include five components, i. e. , AP-1, AP-2, AP-3, AP-3＇ and AP-4. Among these components, AP-3 is an active and potent antitumor maytansinoid, which is also the target fermentation product. The yield and composition of ansamitocins by Actinosynnema mirum were observed to be significantly affected by the addition of branched chain amino acids （BCCAs）. Experiment results showed that the AP-3 yield has doubled with the supplementation of 0.5 g/L valine at 48 h of fermentation, while adding 0.5 g/L leucine or isoleucine exhibited significant inhibition on AP-3 yield （decreased by 58.8% or 83.3G, respectively）. Instead, the contents of AP-2 and AP-4 were increased when leucine or isoleucine was added into the medium. In addition, the secretion of sucelnle that and the side acid and acetic acid was reduced with the addition of valine but increased by isoleucine, implying metabolism of BCCAs in Actinosynnema. mirum influences not only the synthesis of polyketone chains, but also the metabolic state of cells.