Abstract:
Type VII secretion (T7S) systems, also referred to as ESAT-6 secretion (ESX) systems, are molecular machines that have gained great attention due to their implications in cell homeostasis and in host-pathogen interactions in mycobacteria. The latter include important human pathogens such as Mycobacterium tuberculosis (Mtb), the etiological cause of human tuberculosis, which constitutes a pandemic accounting for more than one million deaths every year. The ESX-5 system is exclusively found in slow-growing pathogenic mycobacteria, where it mediates the secretion of a large family of virulence factors: the PE and PPE proteins. The secretion driving force is provided by EccC5, a multidomain ATPase that operates using four globular cytosolic domains: an N-terminal domain of unknown function (EccC5DUF) and three FtsK/SpoIIIE ATPase domains. Recent structural and functional studies of ESX-3 and ESX-5 systems have revealed EccCDUF to be an ATPase-like fold domain with potential ATPase activity, the functionality of which is essential for secretion. Here, the crystal structure of the MtbEccC5DUF domain is reported at 2.05 Å resolution, which reveals a nucleotide-free structure with degenerated cis-acting and trans-acting elements involved in ATP binding and hydrolysis. This crystallographic study, together with a biophysical assessment of the interaction of MtbEccC5DUF with ATP/Mg2+, supports the absence of ATPase activity proposed for this domain. It is shown that this degeneration is also present in DUF domains from other ESX and ESX-like systems, which are likely to exhibit poor or null ATPase activity. Moreover, based on an in silico model of the N-terminal region of MtbEccC5DUF, it is hypothesized that MtbEccC5DUF is a degenerated ATPase domain that may have retained the ability to hexamerize. These observations draw attention to DUF domains as structural elements with potential implications in the opening and closure of the membrane pore during the secretion process via their involvement in inter-protomer interactions.
摘要:
VII型分泌(T7S)系统,也称为ESAT-6分泌(ESX)系统,是分子机器,由于它们在分枝杆菌中的细胞稳态和宿主-病原体相互作用中的影响而获得了极大的关注。后者包括重要的人类病原体,如结核分枝杆菌(Mtb),人类结核病的病因,这构成了每年超过一百万人死亡的大流行。ESX-5系统仅在生长缓慢的致病性分枝杆菌中发现,它介导一大家族毒力因子的分泌:PE和PPE蛋白。分泌驱动力由EccC5提供,EccC5是一种多结构域ATPase,使用四个球形胞质结构域进行操作:一个N末端功能未知的结构域(EccC5DUF)和三个FtsK/SpoIIIEATPase结构域。最近对ESX-3和ESX-5系统的结构和功能研究表明,EccCDUF是具有潜在ATPase活性的ATPase样折叠结构域,其功能对分泌至关重要。这里,MtbEccC5DUF域的晶体结构报告为2.05µ分辨率,它揭示了一个无核苷酸的结构,具有参与ATP结合和水解的简并顺式和反式作用元件。这项晶体学研究,以及对MtbEccC5DUF与ATP/Mg2+相互作用的生物物理评估,支持不存在针对该结构域提出的ATP酶活性。表明,这种变性也存在于其他ESX和ESX样系统的DUF域中,可能表现出不良或无效的ATP酶活性。此外,基于MtbEccC5DUF的N端区域的计算机模拟模型,假设MtbEccC5DUF是一个退化的ATPase结构域,可能保留了六聚体化的能力。这些观察结果引起了人们对DUF结构域作为结构元素的关注,这些结构元素在分泌过程中膜孔的打开和关闭中可能通过参与原聚体间的相互作用。