With the redefinition of polyketide synthase (PKS) modules, a new appreciation of their most downstream domain, the ketosynthase (KS), is emerging. In addition to performing its well-established role of generating a carbon–carbon bond between an acyl-CoA building block and a growing polyketide, it may gatekeep against incompletely processed intermediates. Here, we investigate 739 KSs from 92 primarily actinomycete, cis-acyltransferase assembly lines. When KSs were separated into 16 families based on the chemistries at the α- and β-carbons of their polyketide substrates, a comparison of 32 substrate tunnel residues revealed unique sequence fingerprints. Surprisingly, additional fingerprints were detected when the chemistry at the γ-carbon was considered. Representative KSs were modeled bound to their natural polyketide substrates to better understand observed patterns, such as the substitution of a tryptophan by a smaller residue to accommodate an l-α-methyl group or the substitution of four smaller residues by larger ones to make better contact with a primer unit or diketide. Mutagenesis of a conserved glutamine in a KS within a model triketide synthase indicates that the substrate tunnel is sensitive to alteration and that engineering this KS to accept unnatural substrates may require several mutations.

中文翻译：

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顺式酰基转移酶组装线酮合成酶如何为加工过的聚酮化合物中间体提供把关

随着聚酮合酶 (PKS) 模块的重新定义，对其最下游结构域酮合酶 (KS) 的新认识正在出现。除了发挥其在酰基辅酶 A 结构单元和生长中的聚酮化合物之间产生碳-碳键的公认作用外，它还可以防止未完全加工的中间体。在这里，我们研究了来自 92 种主要放线菌的 739 个 KS，顺式-酰基转移酶装配线。当基于聚酮化合物底物的 α- 和 β- 碳的化学性质将 KS 分成 16 个家族时，对 32 个底物隧道残基的比较揭示了独特的序列指纹。令人惊讶的是，当考虑 γ-碳的化学性质时，检测到了额外的指纹。代表性的 KSs 被建模为与其天然聚酮化合物底物结合，以更好地理解观察到的模式，例如用较小的残基取代色氨酸以适应l-α-甲基或四个较小的残基被较大的残基取代，以更好地与引物单元或二酮化合物接触。模型三酮合成酶中 KS 中保守谷氨酰胺的诱变表明底物隧道对改变敏感，并且设计此 KS 以接受非天然底物可能需要多次突变。