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
生物工程学报 Chin J Biotech 2008, January 25; 24(1): 15-20 Chinese Journal of Biotechnology ISSN 1000-3061 cjb@ ©2008 Institute of Microbiology, CAS & CSM, All rights reserved.
格尔德霉素基因工程高产菌株的构建和培养
赫卫清, 周红霞, 王红远, 高群杰, 王以光
中国医学科学院/中国协和医科大学医药生物技术研究所, 卫生部抗生素生物工程重点实验室, 北京 100050
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ΔSOP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ΔSOP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。 关键词: 吸水链霉菌17997, 格尔德霉素, 基因阻断, 孢子形成周期
Construction and Cultivation of Genetically-engineered Strain to Improve Geldanamycin Production
Weiqing He, Hongxia Zhou, Hongyuan Wang, Qunjie Gao, and Yiguang Wang
Institute of Medicinal Biotechnology, CAMS & PUMC, Key Laboratory of Biotechnology of Antibiotics, Ministry of Health, Beijing 100050, China
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 (ΔSOP) 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 ΔSOP strain increased by 185% comparing with that of the parent strain. On solid medium, the ΔSOP strain underwent 2 cycles of sporulation and the growth of the second sporulation had the highest geldanamycin production.
Keywords: Streptomyces hygroscopicus17997, geldanamycin, gene disruption, spore formation
吸水链霉菌(Streptomyces hygroscopicus)可以产生苯安莎类的Gdm(图1)。安莎类抗生素属于一种大环内酰胺, 它们的结构中都含有一个芳香环(苯醌或萘醌), 其脂肪链是连结在芳香环的两个不相邻原子
之间。芳香环的脂肪链通过酰胺键连接成大环结构。安莎类抗生素芳香环的生物合成均是以AHBA作为起始单元。
安莎类抗生素具有抗菌、抗结核、抗真菌、抗
Received: April 6, 2007; Accepted: June 4, 2007
Supported by: the National Department of Sciences and Technology under Preliminary Basic Research 973 project (No. 2001CCA00500). Corresponding author: Yiguang Wang. Tel: +86-10-63038137; Fax: +86-10-63176489; E-mail: wangyh456@ 科技部基础研究重大项目973前期研究专项(No. 2001CCA00500)资助。