Sweeny Elizabeth A, Jackrel Meredith E, Go Michelle S, Sochor Matthew A, Razzo Beatrice M, DeSantis Morgan E, Gupta Kushol, Shorter James
Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Biochemistry and Molecular Biophysics Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Mol Cell. 2015 Mar 5;57(5):836-849. doi: 10.1016/j.molcel.2014.12.021. Epub 2015 Jan 22.
The structural basis by which Hsp104 dissolves disordered aggregates and prions is unknown. A single subunit within the Hsp104 hexamer can solubilize disordered aggregates, whereas prion dissolution requires collaboration by multiple Hsp104 subunits. Here, we establish that the poorly understood Hsp104 N-terminal domain (NTD) enables this operational plasticity. Hsp104 lacking the NTD (Hsp104(ΔN)) dissolves disordered aggregates but cannot dissolve prions or be potentiated by activating mutations. We define how Hsp104(ΔN) invariably stimulates Sup35 prionogenesis by fragmenting prions without solubilizing Sup35, whereas Hsp104 couples Sup35 prion fragmentation and dissolution. Volumetric reconstruction of Hsp104 hexamers in ATPγS, ADP-AlFx (hydrolysis transition state mimic), and ADP via small-angle X-ray scattering revealed a peristaltic pumping motion upon ATP hydrolysis, which drives directional substrate translocation through the central Hsp104 channel and is profoundly altered in Hsp104(ΔN). We establish that the Hsp104 NTD enables cooperative substrate translocation, which is critical for prion dissolution and potentiated disaggregase activity.
Hsp104溶解无序聚集体和朊病毒的结构基础尚不清楚。Hsp104六聚体中的单个亚基可以溶解无序聚集体，而朊病毒的溶解需要多个Hsp104亚基协同作用。在这里，我们证实了人们了解较少的Hsp104 N端结构域（NTD）赋予了这种操作可塑性。缺失NTD的Hsp104（Hsp104(ΔN)）能够溶解无序聚集体，但无法溶解朊病毒，也不能通过激活突变得到增强。我们确定了Hsp104(ΔN)如何通过切割朊病毒而不溶解Sup35来始终如一地刺激Sup35朊病毒的形成，而Hsp104则将Sup35朊病毒的切割与溶解结合起来。通过小角X射线散射对处于ATPγS、ADP-AlFx（水解过渡态模拟物）和ADP状态下的Hsp104六聚体进行体积重建，揭示了ATP水解时的蠕动泵送运动，该运动驱动底物通过Hsp104中央通道进行定向转运，并且在Hsp104(ΔN)中发生了深刻改变。我们证实，Hsp104 NTD能够实现协同底物转运，这对于朊病毒溶解和增强的解聚酶活性至关重要。
