海绵共生萎缩芽孢杆菌C89 PPTase bap基因的异源表达

Heterologous expression of phosphopantetheinyl transferase gene bap from marine sponge-associated Bacillus atrophaeus C89

摘要:

【背景】磷酸泛酰巯基乙胺基转移酶(PPTase)催化非核糖体肽合成酶(NRPS)中肽酰载体蛋白(PCP)从无活性的脱辅基形态转化为有活性的全辅基形态，从而启动非核糖体肽类化合物的生物合成。【目的】鉴定贪婪倔海绵共生萎缩芽孢杆菌C89中Sfp型PPTase Bap，验证Bap激活NRPS中PCP的能力。【方法】通过BLAST和氨基酸多序列比对鉴定萎缩芽孢杆菌C89中Sfp型PPTase Bap。将bap基因在sfp基因突变株枯草芽孢杆菌168中异源表达，通过重组菌枯草芽孢杆菌168-bap的代谢物检测非核糖体肽类化合物Surfactin。【结果】Bap为Sfp型PPTase，检测到重组菌枯草芽孢杆菌168-bap中Surfactin的产生。【结论】本研究为海洋萎缩芽孢杆菌中NRPS基因簇的异源表达奠定了基础。

关键词:萎缩芽孢杆菌，磷酸泛酰巯基乙胺基转移酶Bap，异源表达

Abstract:

[Background] Phosphopantetheinyl transferases (PPTases) are known to catalyze the transformation of peptidyl carrier proteins (PCPs) in nonribosomal peptide synthetases (NRPSs) from inactive apo-form to active holo-form, so that initiating the synthesis of nonribosomal polypeptides. [Objective] To identify the Sfp-type PPTases Bap from the marine sponge Dysidea avara symbiont Bacillus atrophaeus C89 and to confirm the function of Bap in activating PCPs in NRPSs. [Methods] Sfp-type PPTase Bap from B. atrophaeus C89 was identified by BLAST and amino acid sequences alignment. The bap gene was heterologously expressed in the sfp gene mutant strain Bacillus subtilis 168. The nonribosomal polypeptide surfactin was detected from the metabolites of the recombinant strains B. subtilis 168-bap. [Results] Bap was identified to be Sfp-type PPTases, and the production of surfactin was detected in recombinant strains B. subtilis 168-bap. [Conclusion] Our findings provide basis for heterologous expression of NRPSs gene clusters from marine B. atrophaeus.

Key words:Bacillus atrophaeus, PPTase Bap, Heterologous expression