While only a subset of Vibrio cholerae are human diarrheal pathogens, all are aquatic organisms. In this environment, they often persist in close association with arthropods. In the intestinal lumen of the model arthropod Drosophila melanogaster, methionine and methionine sulfoxide decrease susceptibility to V. cholerae infection. In addition to its structural role in proteins, methionine participates in the methionine cycle, which carries out synthetic and regulatory methylation reactions. It is, therefore, essential for the growth of both animals and bacteria. Methionine is scarce in some environments, and the facile conversion of free methionine to methionine sulfoxide in oxidizing environments interferes with its utilization. To ensure an adequate supply of methionine, the genomes of most organisms encode multiple high affinity uptake pathways for methionine as well as multiple methionine sulfoxide reductases, which reduce free and protein-associated methionine sulfoxide to methionine. To explore the role of methionine uptake and reduction in V. cholerae colonization of the arthropod intestine, we mutagenized the two high affinity methionine transporters and five methionine sulfoxide reductases encoded in the V. cholerae genome. We show that MsrC is the sole methionine sulfoxide reductase active on free methionine sulfoxide. Furthermore, in the absence of methionine synthesis, high affinity methionine uptake but not reduction is essential for V. cholerae colonization of the Drosophila intestine. These findings allow us to place a lower limit of 0.05 mM and an upper limit of 0.5 mM on the methionine concentration in the Drosophila intestine.

Importance Methionine is an essential amino acid involved in both biosynthetic and regulatory processes in the bacterial cell. To ensure an adequate supply of methionine, bacteria have evolved multiple systems to synthesize, import, and recover this amino acid. To explore the importance of methionine synthesis, transport, and recovery in any environment, all these systems must be identified and mutagenized. Here we have mutagenized every high affinity methionine uptake system and methionine sulfoxide reductase encoded in the genome of the diarrheal pathogen V. cholerae. We use this information to determine that high affinity methionine uptake systems are sufficient to acquire methionine in the intestine of the model arthropod Drosophila melanogaster but are not involved in virulence and that the intestinal concentration of methionine must be between 0.05 mM and 0.5 mM.

虽然霍乱弧菌只有一部分是人类腹泻病原体，但它们都是水生生物。在这种环境下，它们经常与节肢动物紧密结合。在节肢动物果蝇模型的肠腔中，蛋氨酸和蛋氨酸亚砜降低了对霍乱弧菌的敏感性感染。蛋氨酸除了在蛋白质中的结构作用外，还参与蛋氨酸循环，该循环进行合成和调节性甲基化反应。因此，它对于动物和细菌的生长都是必不可少的。蛋氨酸在某些环境中是稀缺的，并且在氧化环境中游离蛋氨酸向蛋氨酸亚砜的容易转化会干扰其利用。为了确保充足的蛋氨酸供应，大多数生物体的基因组编码了蛋氨酸的多个高亲和力摄取途径以及多个蛋氨酸亚砜还原酶，这些途径将游离的和蛋白质相关的蛋氨酸亚砜还原为蛋氨酸。探索蛋氨酸摄取和减少在霍乱弧菌中的作用在节肢动物肠道的定殖中，我们诱变了霍乱弧菌基因组中编码的两个高亲和力蛋氨酸转运蛋白和五个蛋氨酸亚砜还原酶。我们表明，MsrC是对游离甲硫氨酸亚砜有活性的唯一甲硫氨酸亚砜还原酶。此外，在不存在蛋氨酸合成的情况下，高亲和力蛋氨酸的摄取而不是减少对于果蝇肠的霍乱弧菌定植是必不可少的。这些发现使我们能够将果蝇肠中蛋氨酸的浓度下限设定为0.05 mM，将上限设定为0.5 mM 。

重要性蛋氨酸是细菌细胞中参与生物合成和调节过程的必需氨基酸。为了确保蛋氨酸的充足供应，细菌已经进化出多种系统来合成，导入和回收该氨基酸。为了探索在任何环境中蛋氨酸合成，运输和回收的重要性，必须对所有这些系统进行识别和诱变。在这里，我们诱变了腹泻病原霍乱弧菌基因组中编码的每个高亲和力蛋氨酸摄取系统和蛋氨酸亚砜还原酶。我们使用此信息来确定高亲和力蛋氨酸摄取系统足以在模型节肢动物果蝇（Drosophila melanogaster）的肠道中获取蛋氨酸。 但不涉及毒力，蛋氨酸的肠道浓度必须在0.05 mM至0.5 mM之间。