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首页> 外文期刊>Biochemistry >Carrier protein recognition in siderophore-producing nonribosomal Peptide synthetases.
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Carrier protein recognition in siderophore-producing nonribosomal Peptide synthetases.

机译：产生铁载体的非核糖体肽合成酶中的载体蛋白识别。

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Nonribosomal peptide synthetases (NRPSs) use phosphopantetheine (pPant) bearing carrier proteins to chaperone activated aminoacyl and peptidyl intermediates to the various enzymes that effect peptide synthesis. Using components from siderophore NRPSs that synthesize vibriobactin, enterobactin, yersiniabactin, pyochelin, and anguibactin, we examined the nature of the interaction of such cofactor-carrier proteins with acyl-activating adenylation (A) domains. While VibE, EntE, and PchD were all able to utilize "carrier protein-free" pPant derivatives, the pattern of usage indicated diversity in the binding mechanism, and even the best substrates were down at least 3 log units relative to the native cofactor-carrier protein. When tested with four noncognate carrier proteins, EntE and VibE differed both in the range of substrate utilization efficiency and in the distribution of the efficiencies across this range. Correlating sequence alignments to kinetic efficiency allowed for the construction of eight point mutants of VibE's worst substrate, HMWP2 ArCP, to the corresponding residue in its native VibB. Mutants S49D and H66E combined to increase activity 6.2-fold and had similar enhancing effects on the downstream condensation domain VibH, indicating that the two NRPS enzymes share carrier protein recognition determinants. Similar mutations of HMWP2 ArCP toward EntB had little effect on EntE, suggesting that the position of recognition determinants varies across NRPS systems.
机译：非核糖体肽合成酶（NRPS）使用带有磷酸泛肽（pPant）的载体蛋白将伴侣蛋白活化的氨酰基和肽基中间体合成到影响肽合成的各种酶上。使用合成荧光弧菌素，肠抑菌素，耶尔西菌素，pyochelin和anguibactin的铁载体NRPSs的成分，我们研究了这种辅因子载体蛋白与酰基激活性腺苷酸化（A）域相互作用的性质。尽管VibE，EntE和PchD都能够利用“不含载体蛋白”的pPant衍生物，但使用方式表明了结合机理的多样性，即使是最佳的底物，相对于天然辅因子-至少下降了3个对数单位。载体蛋白。当用四种非同源载体蛋白进行测试时，EntE和VibE在底物利用效率的范围和效率在该范围内的分布方面都不同。将序列比对与动力学效率相关，可以构建VibE最差底物HMWP2 ArCP的八个点突变体，使其对应于其天然VibB中的相应残基。突变体S49D和H66E结合起来可将活性提高6.2倍，并且对下游的缩合域VibH具有类似的增强作用，表明这两种NRPS酶共享载体蛋白识别决定簇。 HMWP2 ArCP向EntB的相似突变对EntE的影响很小，这表明识别决定因素的位置在NRPS系统中有所不同。

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