The mce1 operon of Mycobacterium tuberculosis, important for lipid metabolism/transport, host cell invasion, modulation of host immune response and pathogenicity, is under the transcriptional control of Mce1R. Hence characterizing Mce1R is an important step for novel anti-tuberculosis drug discovery. The present study reports functional and in silico characterization of Mce1R. In this work, we have computationally modeled the structure of Mce1R and have validated the structure by computational and experimental methods. Mce1R has been shown to harbor the canonical VanR-like structure with a flexible N-terminal 'arm', carrying conserved positively charged residues, most likely involved in the operator DNA binding. The mce1R gene has been cloned, expressed, purified and its DNA-binding activity has been measured in vitro. The K_d value for Mce1R-operator DNA interaction has been determined to be 0.35 ± 0.02 µM which implies that Mce1R binds to DNA with moderate affinity compared to the other FCD family of regulators. So far, this is the first report for measuring the DNA-binding affinity of any VanR-type protein. Despite significant sequence similarity at the N-terminal domain, the wHTH motif of Mce1R exhibits poor conservancy of amino acid residues, critical for DNA-binding, thus results in moderate DNA-binding affinity. The N-terminal DNA-binding domain is structurally dynamic while the C-terminal domain showed significant stability and such profile of structural dynamics is most likely to be preserved in the structural orthologs of Mce1R. In addition to this, a cavity has been detected in the C-terminal domain of Mce1R which contains a few conserved residues. Comparison with other FCD family of regulators suggests that most of the conserved residues might be critical for binding to specific ligand. The max pKd value and drug score for the cavity are estimated to be 9.04 and 109 respectively suggesting that the cavity represents a suitable target site for novel anti-tuberculosis drug discovery approaches.

结核分枝杆菌的mce1操纵子对脂质代谢/运输、宿主细胞侵袭、宿主免疫反应和致病性的调节很重要，受 Mce1R 的转录控制。因此，表征 Mce1R 是发现新型抗结核药物的重要步骤。本研究报告了 Mce1R 的功能和计算机表征。在这项工作中，我们对 Mce1R 的结构进行了计算建模，并通过计算和实验方法验证了该结构。Mce1R 已被证明具有典型的 VanR 样结构，带有灵活的 N 端“臂”，携带保守的带正电残基，最有可能参与操纵基因的 DNA 结合。该mce1R基因已被克隆、表达、纯化，并已在体外测量了其 DNA 结合活性。在K _dMce1R-operator DNA 相互作用的值已确定为 0.35 ± 0.02 µM，这意味着与其他 FCD 调节器家族相比，Mce1R 以中等亲和力与 DNA 结合。到目前为止，这是测量任何 VanR 型蛋白质的 DNA 结合亲和力的第一份报告。尽管 N 端结构域具有显着的序列相似性，但 Mce1R 的 wHTH 基序对氨基酸残基的保守性较差，这对 DNA 结合至关重要，因此导致适度的 DNA 结合亲和力。N 端 DNA 结合域在结构上是动态的，而 C 端域显示出显着的稳定性，这种结构动力学的特征最有可能保留在 Mce1R 的结构直系同源物中。除此之外，在 Mce1R 的 C 端结构域中检测到一个空腔，其中包含一些保守的残基。与其他 FCD 调节剂家族的比较表明，大多数保守残基对于与特定配体的结合可能至关重要。空腔的最大 pKd 值和药物评分估计分别为 9.04 和 109，表明空腔代表了新型抗结核药物发现方法的合适靶点。