School of Biosciences, University of Birmingham, Edgbaston, Birmingham, United Kingdom.
PLoS One. 2011 Mar 31;6(3):e18031. doi: 10.1371/journal.pone.0018031.
Understanding how complex antibiotics are synthesised by their producer bacteria is essential for creation of new families of bioactive compounds. Thiomarinols, produced by marine bacteria belonging to the genus Pseudoalteromonas, are hybrids of two independently active species: the pseudomonic acid mixture, mupirocin, which is used clinically against MRSA, and the pyrrothine core of holomycin.
METHODOLOGY/PRINCIPAL FINDINGS: High throughput DNA sequencing of the complete genome of the producer bacterium revealed a novel 97 kb plasmid, pTML1, consisting almost entirely of two distinct gene clusters. Targeted gene knockouts confirmed the role of these clusters in biosynthesis of the two separate components, pseudomonic acid and the pyrrothine, and identified a putative amide synthetase that joins them together. Feeding mupirocin to a mutant unable to make the endogenous pseudomonic acid created a novel hybrid with the pyrrothine via "mutasynthesis" that allows inhibition of mupirocin-resistant isoleucyl-tRNA synthetase, the mupirocin target. A mutant defective in pyrrothine biosynthesis was also able to incorporate alternative amine substrates.
CONCLUSIONS/SIGNIFICANCE: Plasmid pTML1 provides a paradigm for combining independent antibiotic biosynthetic pathways or using mutasynthesis to develop a new family of hybrid derivatives that may extend the effective use of mupirocin against MRSA.
了解复杂抗生素是如何被其产生菌合成的，对于创造新的生物活性化合物家族至关重要。硫霉素由海洋细菌假交替单胞菌属产生，是两种独立活性物质的混合物：假单胞菌酸混合物，即临床上用于治疗耐甲氧西林金黄色葡萄球菌（MRSA）的莫匹罗星，以及霍洛霉素的吡咯烷核心。
方法/主要发现：对产菌完整基因组进行高通量 DNA 测序，揭示了一个新型的 97 kb 质粒 pTML1，几乎完全由两个不同的基因簇组成。靶向基因敲除证实了这些簇在两种不同成分假单胞菌酸和吡咯烷的生物合成中的作用，并鉴定了一个可能的酰胺合成酶，它将它们连接在一起。向不能产生内源性假单胞菌酸的突变体中添加莫匹罗星，通过“突变合成”产生了一种与吡咯烷的新型杂合体，这种杂合体可以抑制莫匹罗星耐药异亮氨酰-tRNA 合成酶，即莫匹罗星的靶标。一个在吡咯烷生物合成中缺陷的突变体也能够结合替代的胺类底物。
结论/意义：质粒 pTML1 为组合独立的抗生素生物合成途径或利用突变合成来开发新的混合衍生物家族提供了范例，这可能会延长莫匹罗星对耐甲氧西林金黄色葡萄球菌的有效使用。
