This study reveals the genetic and biochemical changes underlying the enhanced hyaluronan (HA) biosynthesis in Streptococcus zooepidemicus. After multiple rounds of atmospheric and room temperature plasma (ARTP) mutagenesis combined with novel bovine serum albumin/cetyltrimethylammonium bromide coupled high-throughput screening assay, the HA yield of the mutant was increased by 42.9% and reached 0.813 g L⁻¹ with a molecular weight of 0.54 × 10⁶ Da within 18 h by shaking flask culture. HA production was increased to 4.56 g L⁻¹ by batch culture in 5-L fermenter. Transcriptome sequencing exhibits that distinct mutants have similar genetic changes. Regulation in direction of metabolic flow into the HA biosynthesis, by enhancing genes responsible for the biosynthesis of HA including hasB, glmU and glmM, weaking downstream gene (nagA and nagB) of UDP-GlcNAc and significantly down-regulating transcription of wall-synthesizing genes, resulting in the accumulation of precursors (UDP-GlcA and UDP-GlcNAc) increased by 39.74% and 119.22%, respectively. These associated regulatory genes may provide control point for engineering of the efficient HA-producing cell factory.

本研究揭示了兽疫链球菌中乙酰透明质酸 (HA) 生物合成增强的遗传和生化变化。经过多轮常压和室温等离子体（ARTP）诱变结合新型牛血清白蛋白/十六烷基三甲基溴化铵偶联高通量筛选试验，突变体的HA产量提高了42.9%，达到0.813 g L −1 ，^具有分子筛 摇瓶培养18 h内质量为0.54×10 ^{6 Da。}HA 产量增加到 4.56 g L ⁻¹在 5 L 发酵罐中分批培养。转录组测序表明不同的突变体具有相似的遗传变化。通过增强负责 HA 生物合成的基因（包括hasB、glmU和glmM）、弱化 UDP-GlcNAc 的下游基因（nagA和nagB）并显着下调壁合成基因的转录，调节 HA 生物合成的代谢流方向，导致前体（UDP-GlcA 和 UDP-GlcNAc）的积累分别增加了 39.74% 和 119.22%。这些相关的调节基因可以为高效的 HA 生产细胞工厂的工程化提供控制点。