AztD, a Periplasmic Zinc Metallochaperone to an ATP-binding Cassette (ABC) Transporter System in Paracoccus denitrificans
AztD, a Periplasmic Zinc Metallochaperone to an ATP-binding Cassette (ABC) Transporter System in Paracoccus denitrificans
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DOI：
10.1074/jbc.m115.684506
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发表时间：
2015-12-11
影响因子：
4.8
通讯作者：
Yukl, Erik T.
中科院分区：
文献类型：
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作者：
Handali, Melody;Roychowdhury, Hridindu;Yukl, Erik T.
过渡金属的细菌atp结合盒（ABC）转运体对于从环境中获取必需元素至关重要。大量革兰氏阴性菌，包括人类病原体，具有第四个功能未知的保守基因，邻近AztABC锌转运体系统的典型渗透酶、atp酶和溶质结合蛋白（SBP）基因。为了评估反硝化副球菌中这种推定的辅助因子（AztD）的功能，我们分析了它的转录调控、金属结合特性以及与收缩压（AztC）的相互作用。锌饥饿条件下aztD基因的转录显著上调。重组表达的AztD从大肠杆菌的周质中获得少量亚化学计量量的锌，能够以高亲和力结合多达三种锌离子。用粒径排除色谱法和简单的本征荧光法确定分离的AztD能够将结合的锌几乎定量地转移到载脂蛋白aztc上。转移是通过一种直接的结合机制发生的，这种机制可以防止金属向溶剂中流失。这些结果表明，AztD是AztC的锌伴侣，可能通过与AztABC系统的相互作用来维持锌的稳态。这项工作扩展了我们对质周锌运输和伴侣蛋白在这一过程中的作用的理解。
Bacterial ATP-binding cassette (ABC) transporters of transition metals are essential for acquisition of necessary elements from the environment. A large number of Gram-negative bacteria, including human pathogens, have a fourth conserved gene of unknown function adjacent to the canonical permease, ATPase, and solute-binding protein (SBP) genes of the AztABC zinc transporter system. To assess the function of this putative accessory factor (AztD) from Paracoccus denitrificans, we have analyzed its transcriptional regulation, metal binding properties, and interaction with the SBP (AztC). Transcription of the aztD gene is significantly up-regulated under conditions of zinc starvation. Recombinantly expressed AztD purifies with slightly substoichiometric zinc from the periplasm of Escherichia coli and is capable of binding up to three zinc ions with high affinity. Size exclusion chromatography and a simple intrinsic fluorescence assay were used to determine that AztD as isolated is able to transfer bound zinc nearly quantitatively to apo-AztC. Transfer occurs through a direct, associative mechanism that prevents loss of metal to the solvent. These results indicate that AztD is a zinc chaperone to AztC and likely functions to maintain zinc homeostasis through interaction with the AztABC system. This work extends our understanding of periplasmic zinc trafficking and the function of chaperones in this process.