L-精氨酸高产菌的选育及基于代谢流量分布的育种机制  

Improvement of high L-arginine producing strain and the mechanism analysis based on metabolic flux distribution

作　　者：许正宏[1] 熊筱晶[1] 窦文芳[1] 陶文沂[1] 

机构地区：[1]江南大学生物工程学院,无锡214036

出　　处：《工业微生物》2006年第4期1-6,共6页Industrial Microbiology

基　　金：国家"十五"科技重点攻关项目(2004BA713B07-05)

摘　　要：从分析钝齿棒杆菌(Corynebacterium crenatum)的精氨酸合成途径入手,提出了一种通过选育脯氨酸结构类似物抗性突变株以提高其精氨酸合成能力的育种思路。采用亚硝基胍(NTG)诱变处理出发菌株YD8(His-,SGr1.2 mg/mL,D-Argr15 mg/mL),经含15 mg/mL的脯氨酸结构类似物S-甲基半胱氨酸(S-MC)的抗性筛选获得精氨酸高产突变株YDM403(His-,SGr1.2 mg/mL,D-Argr15 mg/mL,S-MCr15 mg/mL),产酸水平可达29.4 g/L,较出发菌株YD8的产酸高出55.0%。通过代谢流量分布分析了菌株YD8和YDM403代谢网络的变化,结果表明,菌株YD8可能存在顺序反馈抑制作用,出发株YD8解除了Arg对Glu到Arg的反馈抑制,而YDM403又解除了Pro对Glu到Pro的反馈抑制,从而使中间物Glu累积量下降而对-αKG到Glu不再有反馈抑制,其通量提高,与此同时从Glu向Arg的代谢通量也相应增加。A new approach to screening L-arginine high-producer with the proline analog resistance was proposed by analyzing the L-arginine metabolic pathway of Corynebacteriurn crenaturn. A mutant, YDM403 （His^- , SG^r 1.2 mg/mL, D-Arg^r 15 mg/mL, S-MC15 mg/mL） was selected by the treatment of NTG and praline analog （ S-Methyl Cysteine） resistance using YD8 （His^-, SG^r 1,2 mg/mL, D-Arg^r 15 mg/mL） as a parent strain. Swain YDM403 can yield 29.4g/L of L-arginine, 55.0% higher than that of parent strain YD8. Their metabolic flux distributions of the parent swain YD8 and mutam YDM403 were analyzed. The result showed that strainYD8 depressed the end-prdouct of the enzymes in the pathway from glutamate to arginine. In addition, mutant YDM403 with S-MC resistance depressed the end-product feedback control of the enzymes in the pathway from glutamate to proline, thus depressed the feedback control from α-ketoglutarate to glutamate because the accumulation of glutamate as their precousor was decreased. Furthermore, the synthesis of L-arginine by mutant YDM403 was improved via increasing of the metabolic flux into glutamate.

关 键 词：钝齿棒杆菌 精氨酸 代谢流量分布 菌种选育 

分 类 号：TQ922.9[轻工技术与工程—发酵工程]