透明颤菌血红蛋白基因在红色糖多孢菌株中表达及对红霉素产量的影响_药学论文.docVIP

透明颤菌血红蛋白基因在红色糖多孢菌株中表达及对红霉素产量的影响_药学论文 透明颤菌血红蛋白基因在红色糖多孢菌株中表达及对红霉素产量的影响_药学论文 作者：吴益民 王洪军 孙艳 吴琼 冯立 张志强 杨青 杨国平 【摘要】 目的 利用已含有血红蛋白基因(vgb)的红色糖多孢基因工程重组菌，探讨vgb表达产物对重组糖多孢红霉菌提高红霉素产率的影响。方法 用SDS、Western blot方法鉴定血红蛋白，用葡萄糖浓度与总蛋白质含量比较原始菌株与重组菌株的差别。结果 红色糖多孢基因工程重组菌与原始菌株比较，重组菌株发酵过程葡萄糖浓度的变化出现在第二时段(约38h)，原始菌株达到了0.4g/L，而重组菌株只有0.25g/L；第三时段原始菌株出现了游离葡萄糖浓度高峰而重组菌株未出现。在两个菌株中生物物质浓度［以总蛋白质(g/L)表示］存在不同，重组菌株比原始菌株约低27%。红霉素效价，原始菌株和重组菌株分别为3.98和5.15g/L，相当于重组菌株提高红霉素体积产率约29%。结论 重组红色糖多孢菌表达了透明颤菌血红蛋白，提高了红霉素产率，对解决抗生素工业和基因工程菌高密度发酵有良好的应用前景。 【关键词】 透明颤菌； 血红蛋白基因； 红色糖多孢菌； 基因重组 ABSTRACT Objective To study the exprssion of Vitreoscilla hemoglobin gene (vgb) in the Saccharopoblyspoura erythraea and its effect on erythromycin production. Method A vgb gene was integrated into the chromosomal DNA of Sac.erythraea by electrotransformation and the recombinant Sac.erythraea was obtained. The VHb was identified with SDSand Western blotting method. Glucose concentrations and total protein content in original Sac.erythraea and recombinant strain fermentation processes were determined and compared with auto-biochemical assay. The erythromycin titers were tested with cylinder plate method. Results At the second stage of fermentation, glucose concentration of original strain reached 0.4g/L at about 38h and that of recombinant strain was 0.25g/L; at the third stage, a glucose concentration peak appeared in original strain while none did in recombinant. The total protein content of recombinant was lower than that of original strain by 27%. The titres of erythromycin in original strain and recombinant were 3.98 and 5.15g/L, respectively, i.e. the the erythromycin production of recombinant was increased by 29% compared with that of original strain. Conclusion vgb gene was expressed in recombinant Sac.erythraea, which increased erythromycin production and showed a bright application prospect of antibiotic-gene engineering strain. KEY WORDS Vitreoscilla; vgb; Saccharopolyspora erythraea; Gentic recombination 红霉素是由红色糖多孢菌培养液中分离出来的一种抗生素