格尔德霉素基因工程高产菌株的构建和培养

Construction and Cultivation of Genetically-engineered Strain to Improve Geldanamycin Production

摘要:

在格尔德霉素产生菌吸水链霉菌17997(Streptomyces hygroscopicus 17997)中存在两种3-氨基-5-羟基苯甲酸(3-amino-5-hydroxybenzoic acid, AHBA)的生物合成基因簇, 根据同源性可分为苯醌类和萘醌类。已证明其中苯醌类的AHBA生物合成基因簇负责格尔德霉素(geldanamycin, Gdm)起始单位的合成, 而萘醌类的AHBA基因簇可能参与未知安莎化合物的生物合成。为提高吸水链霉菌17997菌种的Gdm发酵产量, 并研究高产菌种在固体培养基上孢子的生长周期。采用基因阻断技术, 将吸水链霉菌17997中的萘醌类AHBA生物合成基因簇(shnSOP)进行破坏, 以获得DSOP菌株, 从而减少对合成所需共同底物AHBA的争夺。HPLC分析结果表明DSOP菌株Gdm的发酵产量比原株提高185%。同时, 通过孢子计数发现该菌株在固体培养基上的孢子生长经历2个周期, 第2代孢子菌种的Gdm产量较高。

Abstract:

To improve the production of geldanamycin in Streptomyces hygroscopicus 17997, gene disruption was done to delete the naphthalenic AHBA genes (shnSOP), encoding the products that share the common biosynthetic substrates with geldanamycin. The resulting mutant strain (DSOP) was cultivated on a solid medium and the amount of spores collected from the plates was calculated from 5 to 14 days and the yield of geldanamycin was measured by HPLC. The geldanamycin production of the DSOP strain increased by 185% comparing with that of the parent strain. On solid medium, the DSOP strain underwent 2 cycles of sporulation and the growth of the second sporulation had the highest geldanamycin production.