吸水链霉菌井冈变种的色素合成基因缺失对井冈霉素产量的影响

Effects of pigment gene deletions on validamycin A production in Streptomyces hygroscopicus var. jinggangensis

摘要:

[目的] 通过缺失井冈霉素高产菌株TL01中4个典型的色素合成基因簇来考察其对井冈霉素产量、菌体生长和发酵液颜色的影响。[方法] 通过同源重组双交换对4个色素合成基因簇进行逐个同框缺失，HPLC检测突变株井冈霉素产量的变化，qRT-PCR检测突变株中井冈霉素合成基因转录变化，通过称量菌丝体干重来绘制其生长曲线。[结果] 和出发菌株TL01相比，多巴类黑色素基因簇缺失株PY06中井冈霉素的发酵产量由原来的20.6 g/L上升至23.1 g/L，提高了12%；Ⅲ型聚酮合酶编码的黑色素基因簇缺失株PY07产量无明显变化；Ⅱ型聚酮合酶编码的孢子色素基因簇缺失和褐黄素基因簇缺失分别导致井冈霉素产量下降11.7%和17.2%。所有缺失突变株中井冈霉素基因簇转录水平和发酵液颜色均没有明显变化。[结论] 不同色素基因的缺失对井冈霉素产量和菌株生物量积累具有不同的影响。多巴类黑色素与井冈霉素的生物合成过程竞争共同前体，将其中断后使前体流向井冈霉素生物合成，达到了进一步提高产量的目的。

Abstract:

[Objective] We studied the contributions of four pigment biosynthetic genes to validamycin A yield, biomass accumulation, and the color of fermentation broth via individual gene deletions.[Methods] The deletion mutants were obtained via homologous recombination. The titer of validamycin A was detected by HPLC. The transcription of validamycin biosynthetic genes was quantified by qRT-PCR, and the growth was measured with dry cell weight.[Results] Compared with the parent strain, the deletion of DOPA melanin genes increased the validamycin A titer from 20.6 to 23.1 g/L (by 12%), whereas the deletion of type Ⅲ polyketide synthase melanin genes showed no effect. The inactivation of type Ⅱ polyketide synthase spore pigment genes and ochronotic pigment genes decreased validamycin A production by 11.7% and 17.2%, respectively. All these mutant strains had no significant change in transcriptional level and the color of supernatant.[Conclusion] Pigment biosynthetic gene deletions showed different effects on validamycin yield and biomass accumulation, and the deletion of DOPA melanin biosynthetic genes redirected the precursor flux and successfully increased the yield of validamycin A.