链霉菌Streptomyces antibioticus NRRL 8167中Naphthgeranine A生物合成基因簇的分析鉴定

Analysis and identification of the naphthgeranine A biosynthetic gene cluster from Streptomyces antibioticus NRRL 8167

摘要:

【背景】微生物来源的天然产物是小分子药物或药物先导物的重要来源。对链霉菌Streptomyces antibioticus NRRL 8167的基因组分析显示，其包含多个次级代谢产物的生物合成基因簇，具有产生多种新化合物的潜力。【目的】对链霉菌S. antibioticus NRRL 8167中次级代谢产物进行研究，以期发现结构新颖或生物活性独特的化合物，并对相应产物的生物合成基因簇和生物合成途径进行解析。【方法】利用HPLC图谱结合特征性紫外吸收和LC-MS方法，排除S. antibioticus NRRL 8167产生的已知化合物，确定具有特殊紫外吸收的化合物作为挖掘对象，然后利用正、反相硅胶柱色谱、高效液相色谱等技术对次级代谢产物进行分离纯化，分离化合物。利用质谱及核磁共振光谱技术对化合物结构进行解析和鉴定；提取链霉菌S. antibioticus NRRL 8167基因组DNA，利用PacBio测序平台进行基因组测序；利用生物信息学对基因组进行注释，并对合成该化合物的基因簇进行定位分析，推导其生物合成途径。【结果】确定这个化合物是naphthgeranine A，属于聚酮类化合物。全基因组序列分析发现S. antibioticus NRRL 8167基因组含有28个次级代谢产物生物合成基因簇，其中基因簇20可能负责naphthgeranine A的生物合成，并对其生物合成途径进行了推导。【结论】基于紫外吸收光谱和质谱特征，从S. antibioticus NRRL 8167菌株的发酵提取物中分离鉴定了一个聚酮类化合物naphthgeranine A。该菌株的全基因组测序为其生物合成基因簇的鉴定提供了前提，对naphthgeranine A生物合成基因簇和生物合成途径的推测为进一步研究这个化合物的生物合成机制奠定了基础。

Abstract:

[Background] Microbial natural products have become an important source of small molecule drugs and prodrugs in recent years. Genomic analysis shows that Streptomyces antibioticus NRRL 8167 contains a variety of biosynthetic gene clusters of natural products, with the potential to generate multiple new compounds. [Objective] Study on the secondary metabolites in S. antibioticus NRRL 8167 to discover compounds with novel structures or unique biological activities, and further explore the corresponding biosynthetic gene clusters. [Methods] Using HPLC-MS and UV spectrum, we excluded the known compounds produced by S. antibioticus NRRL 8167 and identified an unknown compound with special UV-vis absorption as isolation targets. The secondary metabolites were isolated and purified using normal or reverse phase column chromatography and high-performance liquid chromatography (HPLC). The structures of the compounds were elucidated by using mass spectroscopy (MS) and nuclear magnetic resonance (NMR). The genomic DNA of S. antibioticus NRRL 8167 was extracted for genome sequencing by the PacBio sequencing platform. Bioinformatics technique was used to annotate the genome, to define the gene clusters that is responsible for the compound biosynthesis, and to propose its biosynthetic pathway. [Results] The compound was purified and identified as naphthgeranine A, belonging to polyketides. The whole genome sequence contains 28 secondary metabolites biosynthetic gene clusters. The gene cluster 20 is proposed to be responsible for the biosynthesis of naphthgeranine A. We also speculated the biosynthetic pathway of naphthgeranine A. [Conclusion] Naphthgeranine A was purified from S. antibioticus NRRL 8167 and structurally elucidated based on the UV-vis spectra, MS data and NMR. Complete genome sequence of this strain provided the opportunity for defining the gene clusters of naphthgeranine A. The proposed biosynthetic gene cluster and biosynthetic pathways laid the foundation for further research on the biosynthetic mechanisms of naphthgeranine A.