Tripathi Prajna, Parijat Priyanka, Patel Virendra Kumar, Batra Janendra K
National Institute of Immunology New Delhi India.
Present address: Randall Division of Cell and Molecular Biophysics King's College London UK.
FEBS Open Bio. 2018 Sep 15;8(10):1669-1690. doi: 10.1002/2211-5463.12509. eCollection 2018 Oct.
() is known to persist in extremely hostile environments within host macrophages. The ability to withstand such proteotoxic stress comes from its highly conserved molecular chaperone machinery. ClpB, a unique member of the AAA+ family of chaperones, is responsible for resolving aggregates in and many other bacterial pathogens. produces two isoforms of ClpB, a full length and an N-terminally truncated form (ClpB∆N), with the latter arising from an internal translation initiation site. It is not clear why this internal start site is conserved and what role the N-terminal domain (NTD) of ClpB plays in its function. In the current study, we functionally characterized and compared the two isoforms of ClpB. We found the NTD to be dispensable for oligomerization, ATPase activity and prevention of aggregation activity of ClpB. Both ClpB and ClpB∆N were found to be capable of resolubilizing protein aggregates. However, the efficiency of ClpB∆N at resolubilizing higher order aggregates was significantly lower than that of ClpB. Further, ClpB∆N exhibited reduced affinity for substrates as compared to ClpB. We also demonstrated that the surface of the NTD of ClpB has a hydrophobic groove that contains four hydrophobic residues: L97, L101, F140 and V141. These residues act as initial contacts for the substrate and are crucial for stable interaction between ClpB and highly aggregated substrates.
已知（某物质）在宿主巨噬细胞内极其恶劣的环境中持续存在。其耐受这种蛋白毒性应激的能力源于其高度保守的分子伴侣机制。ClpB是AAA + 家族伴侣蛋白中的独特成员，负责分解（某物质）及许多其他细菌病原体中的聚集体。（某物质）产生两种ClpB同工型，一种全长型和一种N端截短型（ClpB∆N），后者源自内部翻译起始位点。目前尚不清楚为何这个内部起始位点是保守的，以及ClpB的N端结构域（NTD）在其功能中起什么作用。在本研究中，我们对ClpB的两种同工型进行了功能表征和比较。我们发现NTD对于ClpB的寡聚化、ATP酶活性和防止聚集活性是可有可无的。发现ClpB和ClpB∆N都能够使蛋白质聚集体重新溶解。然而，ClpB∆N重新溶解高阶聚集体的效率明显低于ClpB。此外，与ClpB相比，ClpB∆N对底物的亲和力降低。我们还证明，ClpB的NTD表面有一个疏水凹槽，其中包含四个疏水残基：L97、L101、F140和V141。这些残基作为底物的初始接触点，对于ClpB与高度聚集的底物之间的稳定相互作用至关重要。
