Characterization of the pyoluteorin biosynthetic gene cluster of Pseudomonas fluorescens Pf-5
Characterization of the pyoluteorin biosynthetic gene cluster of Pseudomonas fluorescens Pf-5
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DOI：
10.1128/jb.181.7.2166-2174.1999
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发表时间：
1999-04-01
影响因子：
3.2
通讯作者：
Loper, JE
中科院分区：
文献类型：
--
作者：
Nowak-Thompson, B;Chaney, N;Loper, JE
在荧光假单胞菌Pf-5的24-kb基因组区域内鉴定了10个pit基因，其中8个pit基因的氨基酸序列与已知生物合成功能基因（包括Ⅰ型聚酮酶）的氨基酸序列相似在一个实施方案中，所述酶包括酰基辅酶A（pltB，pltC）、酰基辅酶A（酰基-CoA）脱氢酶（pltE）、酰基-CoA合成酶（pltF）、硫酯酶（pltG）和三种卤代酶（pltA，pltD和pltM）。转座子Tn 5或Tn 3-nice或卡那霉素抗性基因在这些基因中的每一个中的插入消除了Pf-5在生产中的pyoluences。推测的八种ph产物的功能与参与由脯氨酸和乙酸前体生物合成pyoluences的生化转化一致。同位素标记的研究表明，脯氨酸是二氯吡咯部分的pyolurespecins的主要前体。推导的氨基酸序列的另一个坑基因，pltR的产品，是类似的转录激活因子的LysR家族的成员。pltR和pltM从pltLABCDEFG基因簇中分化转录，并且在分离pltRM和pltLABCDEFG的486-bp基因间区域内鉴定出具有LysR结合位点特征的序列。通过与冰核报告基因的转录融合，评估了Pf-5中的细菌合成基因pltB、pltE和pltF的转录，其显著高于Pf-5的pltR突变体。因此，PltR被认为是生物合成基因中的连锁细菌合成基因的转录激活因子。
Ten genes (plt) required for the biosynthesis of pyoluteorin, an antifungal compound composed of a bichlorinated pyrrole linked to a resorcinol moiety, were identified within a 24-kb genomic region of Pseudomonas fluorescens Pf-5, The deduced amino acid sequences of eight pit genes were similar to the amino acid sequences of genes with known biosynthetic functions, including type I polyketide synthases (pltB, pltC), an acyl coenzyme A (acyl-CoA) dehydrogenase (pltE), an acyl-CoA synthetase (pltF), a thioesterase (pltG), and three halogenases (pltA, pltD, and pltM). Insertions of the transposon Tn5 or Tn3-nice or a kanamycin resistance gene in each of these genes abolished pyoluteorin production by Pf-5, The presumed functions of the eight ph products are consistent with biochemical transformations involved in pyoluteorin biosynthesis from proline and acetate precursors. Isotope labeling studies demonstrated that proline is the primary precursor to the dichloropyrrole moiety of pyoluteorin. The deduced amino acid sequence of the product of another pit gene, pltR, is similar to those of members of the LysR family of transcriptional activators. pltR and pltM are transcribed divergently from the pltLABCDEFG gene cluster, and a sequence with the characteristics of a LysR binding site was identified within the 486-bp intergenic region separating pltRM from pltLABCDEFG. Transcription of the pyoluteorin biosynthesis genes pltB, pltE, and pltF, assessed with transcriptional fusions to an ice nucleation reporter gene, was significantly greater in Pf-5 than in a pltR mutant of Pf-5, Therefore, PltR is proposed to be a transcriptional activator of linked pyoluteorin biosynthesis genes.