摘要
在必特螺旋霉素基因工程链霉菌WSJ～1—195发酵生产必特螺旋霉素的摇瓶实验中，发酵36h时向合成培养基中添加0．5g／L的缬氨酸对必特螺旋霉素效价和组分的影响最为显著。实验结果表明：添加缬氨酸后，菌体在发酵48～60h糖耗加快，达到1g／（L·h），发酵液中丙酸和丁酸浓度先增后降，而发酵液中丙酮酸在胞内积累，丙酮酸羧化酶活性在48h和60h时分别增加20％和95％。由于糖耗加快，更多碳流可以通过甲基丙二酰CoA转羧基酶途径（MCT途径）将缬氨酸分解生成的丙酰CoA和丁酰CoA转化为内酯环合成所需要的直接三碳前体，最终效价比对照高出45．3％。添加缬氨酸后，由于大环合成增多，侧链前体的供应无法满足内酯环增加的需求，导致酰化组分含量总体下降，其中总异戊酰组分降低23％，乙酰螺旋霉素Ⅲ降低30．8％，丙酰螺旋霉素Ⅱ降低33％，丙酰螺旋霉素Ⅲ降低48．8％。同时缬氨酸代谢生成的丁酸也有可能会影响异戊酰基转移酶的活性。
It was found that there is the greatest influence on the biosynthesis of biotechspiramycin by adding 0.5 g/L valine to the chemically defined culture at 36 h when fermentation. The parameters in the cultivation process, such as biological titer, components of biotechspiramycin, rate of sugar consumption, organic acids of extracellular, pyruvate carboxylase （PC） activity were determined. Results show that after supplementation of valine, sugar is used more quickly than the control before 60 h, and the rate of sugar consumption increased to 1g/（L·h）. Pyruvic acid is accumulated in the cell. The activity of pyrurate carboxylase responsible for transforming pyruvate to oxaloacetate increase by 20% and 95% at 48 h and 60 h, respectively. More carbon source is transformed via methylmalonyl-CoA carboxytransferase and propionyl-CoA carboxylaxe paths to malonyl-CoA, methylmalonyl-CoA, ethylmalonyl-CoA, serving as direct precursors for macrolide biosythesis of biotechspiramycin, leading to the increase of biological titers. Due to deficiency of acyl-precursors for the acylation in 4＂ site of biotechspiramycin after adding valine, the proportion of total isovaleryl-spiramycin, acetyl-spiramycin Ⅲ, propionyl-spiramycin Ⅱand propionyl-spi
ramycin Ⅲ are decreased by 23% ,30.8%.33%.48.8%, respectively, but butyl-spiramycins Ⅱ and total spiramycins are increased greatly, and isovaleryl-CoA transferase may be negatively modulated by butyric acid derived from valine.
出处
《华东理工大学学报（自然科学版）》
EI
CAS
CSCD
北大核心
2008年第1期53-59,共7页
Journal of East China University of Science and Technology
基金
上海市科委项目(064319033)
教育部回国留学人员基金
