Department of Microbiology and Molecular Genetics, University of Vermont College of Medicine, 95 Carrigan Drive, 322 Stafford Hall, Burlington, VT 05405, USA.
Appl Environ Microbiol. 2011 Jul;77(13):4383-9. doi: 10.1128/AEM.00504-11. Epub 2011 May 20.
Choline is abundant in association with eukaryotes and plays roles in osmoprotection, thermoprotection, and membrane biosynthesis in many bacteria. Aerobic catabolism of choline is widespread among soil proteobacteria, particularly those associated with eukaryotes. Catabolism of choline as a carbon, nitrogen, and/or energy source may play important roles in association with eukaryotes, including pathogenesis, symbioses, and nutrient cycling. We sought to generate choline analogues to study bacterial choline catabolism in vitro and in situ. Here we report the characterization of a choline analogue, propargylcholine, which inhibits choline catabolism at the level of Dgc enzyme-catalyzed dimethylglycine demethylation in Pseudomonas aeruginosa. We used genetic analyses and 13C nuclear magnetic resonance to demonstrate that propargylcholine is catabolized to its inhibitory form, propargylmethylglycine. Chemically synthesized propargylmethylglycine was also an inhibitor of growth on choline. Bioinformatic analysis suggests that there are genes encoding DgcA homologues in a variety of proteobacteria. We examined the broader utility of propargylcholine and propargylmethylglycine by assessing growth of other members of the proteobacteria that are known to grow on choline and possess putative DgcA homologues. Propargylcholine showed utility as a growth inhibitor in P. aeruginosa but did not inhibit growth in other proteobacteria tested. In contrast, propargylmethylglycine was able to inhibit choline-dependent growth in all tested proteobacteria, including Pseudomonas mendocina, Pseudomonas fluorescens, Pseudomonas putida, Burkholderia cepacia, Burkholderia ambifaria, and Sinorhizobium meliloti. We predict that chemical inhibitors of choline catabolism will be useful for studying this pathway in clinical and environmental isolates and could be a useful tool to study proteobacterial choline catabolism in situ.
胆碱与真核生物密切相关，在许多细菌中发挥渗透保护、热保护和膜生物合成的作用。胆碱的有氧分解在土壤变形杆菌中广泛存在，尤其是那些与真核生物相关的细菌。作为碳、氮和/或能源的胆碱分解代谢可能在与真核生物的共生关系中发挥重要作用，包括发病机制、共生和营养循环。我们试图生成胆碱类似物，以研究细菌胆碱分解代谢的体外和原位情况。在这里，我们报告了一种胆碱类似物，炔丙基胆碱的特征，它在铜绿假单胞菌中抑制 Dgc 酶催化的二甲基甘氨酸脱甲基化的胆碱分解代谢。我们使用遗传分析和 13C 核磁共振证明炔丙基胆碱被代谢为其抑制形式炔丙基甲基甘氨酸。化学合成的炔丙基甲基甘氨酸也是生长在胆碱上的抑制剂。生物信息学分析表明，各种变形杆菌中都有编码 DgcA 同源物的基因。我们通过评估已知在胆碱上生长并具有推定的 DgcA 同源物的其他变形杆菌成员的生长情况，检查了炔丙基胆碱和炔丙基甲基甘氨酸的更广泛用途。炔丙基胆碱可作为铜绿假单胞菌生长的抑制剂，但不能抑制测试的其他变形杆菌的生长。相比之下，炔丙基甲基甘氨酸能够抑制所有测试的变形杆菌，包括门多萨假单胞菌、荧光假单胞菌、恶臭假单胞菌、洋葱伯克霍尔德菌、ambifaria 伯克霍尔德菌和苜蓿中华根瘤菌，依赖胆碱的生长。我们预测，胆碱分解代谢的化学抑制剂将有助于研究临床和环境分离株中的这条途径，并且可能是研究原位变形杆菌胆碱分解代谢的有用工具。
