Department of Bioscience and Biotechnology, Konkuk University, Seoul, 05029, Republic of Korea.
Sci Rep. 2021 Apr 12;11(1):7945. doi: 10.1038/s41598-021-86997-3.
Some Gram-negative bacteria harbor lipids with aryl polyene (APE) moieties. Biosynthesis gene clusters (BGCs) for APE biosynthesis exhibit striking similarities with fatty acid synthase (FAS) genes. Despite their broad distribution among pathogenic and symbiotic bacteria, the detailed roles of the metabolic products of APE gene clusters are unclear. Here, we determined the crystal structures of the β-ketoacyl-acyl carrier protein (ACP) reductase ApeQ produced by an APE gene cluster from clinically isolated virulent Acinetobacter baumannii in two states (bound and unbound to NADPH). An in vitro visible absorption spectrum assay of the APE polyene moiety revealed that the β-ketoacyl-ACP reductase FabG from the A. baumannii FAS gene cluster cannot be substituted for ApeQ in APE biosynthesis. Comparison with the FabG structure exhibited distinct surface electrostatic potential profiles for ApeQ, suggesting a positively charged arginine patch as the cognate ACP-binding site. Binding modeling for the aryl group predicted that Leu185 (Phe183 in FabG) in ApeQ is responsible for 4-benzoyl moiety recognition. Isothermal titration and arginine patch mutagenesis experiments corroborated these results. These structure-function insights of a unique reductase in the APE BGC in comparison with FAS provide new directions for elucidating host-pathogen interaction mechanisms and novel antibiotics discovery.
一些革兰氏阴性菌含有带有芳基聚烯（APE）部分的脂质。APE 生物合成的生物合成基因簇（BGCs）与脂肪酸合酶（FAS）基因表现出惊人的相似性。尽管它们在致病和共生细菌中广泛分布，但 APE 基因簇代谢产物的详细作用尚不清楚。在这里，我们确定了临床分离的毒力鲍曼不动杆菌 APE 基因簇产生的β-酮酰-酰基载体蛋白（ACP）还原酶 ApeQ 在两种状态（与 NADPH 结合和未结合）下的晶体结构。APE 聚烯部分的体外可见吸收光谱测定表明，来自 A.baumannii FAS 基因簇的β-酮酰-ACP 还原酶 FabG 不能替代 APE 生物合成中的 ApeQ。与 FabG 结构的比较显示 ApeQ 具有独特的表面静电势分布，表明正电荷精氨酸斑是 ACP 的结合位点。芳基的结合建模预测 ApeQ 中的 Leu185（FabG 中的 Phe183）负责识别 4-苯甲酰部分。等温滴定和精氨酸斑诱变实验证实了这些结果。与 FAS 相比，APE BGC 中这种独特还原酶的结构-功能研究为阐明宿主-病原体相互作用机制和发现新型抗生素提供了新的方向。
