Background

Mycobacterial FASII pathway is governed by the Protein-Protein Interaction mediated dynamics existent between Acyl Carrier Protein and its partner enzymes. The dehydratase HadAB, involved in the third step of FASII synthesis has remained a key target of drugs like Thiacetazone (TAC) and its consequence on AcpM binding is yet to be deciphered. Owing to the transient nature of these interactions, analysing their implications as a drug target has been exhausting.

Methods

In this context, we have developed an in vitro method to study the effect of thiocarbamide-containing compounds, TAC and SPA0355 (a thiourea analogue) against mycobacterial HadAB. Additionally, by utilizing crypto-ACP (NBD-tagged Acyl Carrier Protein) as a tool of our choice, we attempted at exploring the effect of TAC and SPA0355 on mycobacterial HadAB.

Results

SPA0355 behaves at par with TAC and undergoes activation in the presence of monooxygenase EthA thus, bringing about a covalent modification in HadA subunit of HadAB. The crypto-ACP method provides insights into the altered substrate housing capability in HadAB associated with the impediment of its AcpM mediated functionality; an outcome attributed to the repercussions associated with the binding of the aforementioned thiourea compounds.

Conclusion

This investigation has assisted in unveiling a two-step mechanism undertaken by AcpM for interacting with its corresponding partner protein during acyl chain transfer.

General significance

This study highlights the alterations brought about by drug binding in the interplay between ACP and HadAB. Additionally, this work for the first time establishes the role of SPA0355 as a promising drug candidate against dehydratase HadAB.

背景

分枝杆菌 FASII 途径受蛋白质-蛋白质相互作用介导的酰基载体蛋白与其伙伴酶之间存在的动力学控制。参与 FASII 合成第三步的脱水酶 HadAB 仍然是噻乙酰酮 (TAC) 等药物的关键目标，其对 AcpM 结合的影响尚待破译。由于这些相互作用的短暂性，分析它们作为药物靶标的含义已经很费劲了。

方法

在这种情况下，我们开发了一种体外方法来研究含硫脲化合物、TAC 和 SPA0355（一种硫脲类似物）对分枝杆菌 HadAB 的影响。此外，通过使用加密 ACP（带有 NBD 标签的酰基载体蛋白）作为我们选择的工具，我们尝试探索 TAC 和 SPA0355 对分枝杆菌 HadAB 的影响。

结果

SPA0355 的行为与 TAC 相同，并在单加氧酶 EthA 存在下进行活化，从而导致 HadAB 的 HadA 亚基发生共价修饰。加密 ACP 方法提供了对与 AcpM 介导的功能障碍相关的 HadAB 中改变的基板外壳能力的见解；结果归因于与上述硫脲化合物的结合相关的影响。

结论

这项研究有助于揭示 AcpM 在酰基链转移过程中与其相应伙伴蛋白相互作用的两步机制。

一般意义

这项研究强调了药物结合在 ACP 和 HadAB 之间的相互作用中带来的改变。此外，这项工作首次确立了 SPA0355 作为一种有前景的抗脱水酶 HadAB 候选药物的作用。