常压室温等离子体诱变高产去甲金霉素金色链霉菌

Mutation of Streptomyces aureofaciens with High Yield Demethylchlortetracycline by Atmospheric and Room Temperature Plasma

采用新型常压室温等离子体(ARTP)诱变技术,基于96孔板高通量筛选及高效液相色谱检测,获得一株金色链霉菌突变株A6-6-9,其去甲金霉素(1)产量较出发菌提高22.5%。经4次传代,突变株A6-6-9的遗传稳定性良好。与出发菌相比,在相同的摇瓶发酵过程中,突变株氨基氮利用率加快,生长和糖消耗速率减慢,产1能力提高。有机酸的变化趋势表明,突变株比出发菌积累更多的草酰乙酸、丙二酸等前体有机酸,流向丙二酰辅酶A的合成,进而提高1产量。二维电泳及质谱结果显示,苹果酸脱氢酶、谷氨酰胺合成酶、6-羟基化酶等与1合成密切相关的酶在突变株中表达更多,推测也是突变株高产1的一个重要原因。

Atmospheric and room temperature plasma （ARTP） mutation technique was used and combined with high-throughput screening based on 96 well plates and HPLC to obtain Streptomyces aureofaciens mutants with high demethylchlortetracycline （1） productivity.A mutant A6-6-9 was screened and showed high genetic stability after four subcultures.Compared with the original strain under the same flask fermentation process,the mutant A6-6-9 showed faster aminonitrogen consumption,slower growth rate and glucose consumption rate,and enhanced productivity of 1 with an increase of 22.5 ％.The changes of organic acids were also studied.Compared with the original strain,the mutant accumulated more precursor organic acids such as oxaloacetic acid and malonic acid which entered into the synthesis of malonyl-CoA,finally enhanced the production of 1.2D electrophoresis （2-DE） and protein points by MS showed that malate dehydrogenase,glutamine synthetase and 6-hydroxylation enzyme expressed more in the mutant strain,which might be an important reason for the enhancement of 1 productivity.