Chen S, Roberts J B, Xue Y, Sherman D H, Reynolds K A
Department of Medicinal Chemistry and Institute for Structural Biology and Drug Discovery, Virginia Commonwealth University, 800 East Leigh Street, Richmond, VA 23219, USA.
Gene. 2001 Jan 24;263(1-2):255-64. doi: 10.1016/s0378-1119(00)00560-6.
In Streptomyces venezuelae, four polyketide synthase (PKS) polypeptides encoded by pikAI-pikAIV are used to generate 10 and 12-membered macrocyclic structures, narbonolide and 10-deoxymethynolide. Sequence analysis suggests these genes are translationally coupled with downstream genes, pikAV (encoding a type II thioesterase), desVIII-desVI (encoding enzymes responsible for production of the final glycosylated products pikromycin, narbomycin, methymycin and neomethymycin) and desR (a resistance gene). Type II thioesterases have been suggested to have an editing function in polyketide biosynthesis and deletion of the corresponding genes often leads to decreased levels of polyketide production. Surprisingly an in-frame deletion of 687 bp of the 843 bp pikAV ORF led to a strain SC1022 that produced normal yields of polyketide products, but only in the aglycone form. Plasmid-based expression of the desVIII-VI and desR in the SC1022 strain completely restored production of glycosylated products, despite the absence of a functional pikAV gene product. Under these conditions the PikAV TEII therefore does not play an important role in polyketide biosynthesis, and its function remains an enigma. These observations also demonstrate that the region of pikAV DNA deleted in strain SC1022 contains a transcription unit essential for expression of the des genes. A sequence alignment of PikAV with members of the highly conserved type II thioesterases revealed a short divergent region at the carboxy terminus, suggesting a region of pikAV that might contain such a transcription unit. DNA containing this region of pikAV was shown to be able to increase plasmid-based expression of both crotonyl CoA reductase gene (ccr) and the erythromycin resistance gene (ermE) in S. venezuelae.
在委内瑞拉链霉菌中，由pikAI - pikAIV编码的四种聚酮合酶（PKS）多肽用于生成10元和12元大环结构，即纳波内酯和10 - 脱氧甲炔诺内酯。序列分析表明，这些基因与下游基因翻译偶联，pikAV（编码II型硫酯酶）、desVIII - desVI（编码负责最终糖基化产物匹克霉素、纳波霉素、甲基霉素和新甲基霉素生成的酶）和desR（一个抗性基因）。II型硫酯酶被认为在聚酮生物合成中具有编辑功能，相应基因的缺失通常会导致聚酮产量降低。令人惊讶的是，843 bp的pikAV开放阅读框（ORF）内687 bp的框内缺失产生了菌株SC1022，该菌株聚酮产物产量正常，但仅以苷元形式存在。尽管缺乏功能性的pikAV基因产物，但在SC1022菌株中基于质粒表达desVIII - VI和desR完全恢复了糖基化产物的产生。因此，在这些条件下，PikAV TEII在聚酮生物合成中不发挥重要作用，其功能仍是个谜。这些观察结果还表明，菌株SC1022中缺失的pikAV DNA区域包含一个对des基因表达至关重要的转录单元。PikAV与高度保守的II型硫酯酶成员的序列比对揭示了羧基末端的一个短的差异区域，表明pikAV的一个区域可能包含这样一个转录单元。含有pikAV该区域的DNA能够增加委内瑞拉链霉菌中巴豆酰辅酶A还原酶基因（ccr）和红霉素抗性基因（ermE）基于质粒的表达。
