Structural, Biochemical, and In Vivo Characterization of MtrR-Mediated Resistance to Innate Antimicrobials by the Human Pathogen Neisseria gonorrhoeae
Structural, Biochemical, and In Vivo Characterization of MtrR-Mediated Resistance to Innate Antimicrobials by the Human Pathogen Neisseria gonorrhoeae
复制标题
DOI：
10.1128/jb.00401-19
复制
发表时间：
2019-10-01
影响因子：
3.2
通讯作者：
Brennan, Richard G.
中科院分区：
文献类型：
--
作者：
Beggs, Grace A.;Zalucki, Yaramah M.;Brennan, Richard G.
淋病奈瑟菌通过诱导 mtrCDE 编码的多药物外排泵的表达来响应宿主来源的抗菌剂，该泵将杀微生物剂（例如胆汁盐、脂肪酸和多种外源性药物）从细胞中排出。在没有这些细胞毒素的情况下，TetR 家族成员 MtrR 会抑制 mtrCDE 基因。尽管抗菌药物依赖性 mtrCDE 去抑制是明确的，但 MtrR 的生理诱导剂尚不清楚。在这里，我们报告了 MtrR 诱导形式的晶体结构。在 MtrR 的结合口袋中，我们观察到电子密度，我们假设是 N-环己基-3-氨基丙磺酸 (CAPS)，结晶试剂的一种成分。使用 MtrR-CAPS 结构作为诱导剂结合模板，我们假设胆汁盐与 CAPS 具有显着的化学相似性，是生理相关的诱导剂。事实上，MtrR-鹅脱氧胆酸盐和MtrR-牛磺脱氧胆酸盐相互作用的体外和体内表征表明，这些胆汁盐（而不是甘胆酸盐或牛磺胆酸盐）与MtrR紧密结合，并且可以充当真正的诱导剂。此外，W136 和 R176 这两个残基被证明对结合鹅脱氧胆酸盐很重要，但对牛磺脱氧胆酸盐则不然，这表明胆汁盐的不同结合模式。这些数据提供了对病原体用来克服人类先天防御的关键机制的深入了解。 重要性 淋病奈瑟菌在全世界范围内造成了重大的疾病负担，其多药耐药性的急剧上升降低了以前或目前批准用于治疗淋病感染的抗生素的功效。多药外排泵 MtrCDE 从细菌细胞中运输多种药物和宿主来源的抗菌剂，并赋予宿主内病原体的生存优势。泵的转录受到 MtrR 的抑制，但通过胞质内抗生素的流入而缓解。在这里，我们描述了诱导的 MtrR 的结构，并利用该结构来鉴定胆汁盐作为 MtrR 的生理诱导剂。这些发现为 MtrR 在暴露于内在和临床应用的抗菌药物后通过解除 mtrCDE 表达的抑制，为抗菌传感和淋球菌保护提供了机制基础。
Neisseria gonorrhoeae responds to host-derived antimicrobials by inducing the expression of the mtrCDE-encoded multidrug efflux pump, which expels microbicides, such as bile salts, fatty acids, and multiple extrinsically administered drugs, from the cell. In the absence of these cytotoxins, the TetR family member MtrR represses the mtrCDE genes. Although antimicrobial-dependent derepression of mtrCDE is clear, the physiological inducers of MtrR are unknown. Here, we report the crystal structure of an induced form of MtrR. In the binding pocket of MtrR, we observed electron density that we hypothesized was N-cyclohexyl-3-aminopropanesulfonic acid (CAPS), a component of the crystallization reagent. Using the MtrR-CAPS structure as an inducer-bound template, we hypothesized that bile salts, which bear significant chemical resemblance to CAPS, are physiologically relevant inducers. Indeed, characterization of MtrR-chenodeoxycholate and MtrR-taurodeoxycholate interactions, both in vitro and in vivo, revealed that these bile salts, but not glyocholate or taurocholate, bind MtrR tightly and can act as bona fide inducers. Furthermore, two residues, W136 and R176, were shown to be important in binding chenodeoxycholate but not taurodeoxycholate, suggesting different binding modes of the bile salts. These data provide insight into a crucial mechanism utilized by the pathogen to overcome innate human defenses.IMPORTANCE Neisseria gonorrhoeae causes a significant disease burden worldwide, and a meteoric rise in its multidrug resistance has reduced the efficacy of antibiotics previously or currently approved for therapy of gonorrheal infections. The multidrug efflux pump MtrCDE transports multiple drugs and host-derived antimicrobials from the bacterial cell and confers survival advantage on the pathogen within the host. Transcription of the pump is repressed by MtrR but relieved by the cytosolic influx of antimicrobials. Here, we describe the structure of induced MtrR and use this structure to identify bile salts as physiological inducers of MtrR. These findings provide a mechanistic basis for antimicrobial sensing and gonococcal protection by MtrR through the derepression of mtrCDE expression after exposure to intrinsic and clinically applied antimicrobials.