Beuron F, Maurizi M R, Belnap D M, Kocsis E, Booy F P, Kessel M, Steven A C
Laboratory of Structural Biology, National Institute of Arthritis, Musculoskeletal and Skin Diseases, Bethesda, Maryland, 20892, USA.
J Struct Biol. 1998 Nov;123(3):248-59. doi: 10.1006/jsbi.1998.4039.
ClpAP, a typical energy-dependent protease, consists of a proteolytic component (ClpP) and a chaperone-like ATPase (ClpA). ClpP is composed of two apposed heptameric rings, whereas in the presence of ATP or ATPgammaS, ClpA is a single hexameric ring. Formation of ClpAP complexes involves a symmetry mismatch as sixfold ClpA stacks axially on one or both faces of sevenfold ClpP. We have analyzed these structures by cryo-electron microscopy. Our three-dimensional reconstruction of ClpA at 29-A resolution shows the monomer to be composed of two domains of similar size that, in the hexamer, form two tiers enclosing a large cavity. Cylindrical reconstruction of ClpAP reveals three compartments: the digestion chamber inside ClpP; a compartment between ClpP and ClpA; and the cavity inside ClpA. They are connected axially via narrow apertures, implying that substrate proteins should be unfolded to allow translocation into the digestion chamber. The cavity inside ClpA is structurally comparable to the "Anfinsen cage" of other chaperones and may play a role in the unfolding of substrates. A geometrical description of the symmetry mismatch was obtained by using our model of ClpA and the crystal structure of ClpP (Wang et al., 1997, Cell 91, 447-456) to identify the particular side views presented by both molecules in individual complexes. The interaction is characterized by a key pair of subunits, one of each protein. A small turn (8.6(o) = 2pi/42; equivalent to a 4-A shift) would transfer the key interaction to another pair of subunits. We propose that nucleotide hydrolysis results in rotation, facilitating the processive digestion of substrate proteins.
ClpAP是一种典型的能量依赖性蛋白酶，由一个蛋白水解成分（ClpP）和一个类似伴侣的ATP酶（ClpA）组成。ClpP由两个相对的七聚体环组成，而在ATP或ATPγS存在的情况下，ClpA是一个单一的六聚体环。ClpAP复合物的形成涉及对称性不匹配，因为六聚体的ClpA在七聚体的ClpP的一个或两个面上轴向堆叠。我们通过冷冻电子显微镜分析了这些结构。我们以29埃分辨率对ClpA进行的三维重建显示，单体由两个大小相似的结构域组成，在六聚体中，这两个结构域形成两层，包围着一个大腔。ClpAP的圆柱形重建揭示了三个隔室：ClpP内部的消化室；ClpP和ClpA之间的隔室；以及ClpA内部的腔。它们通过狭窄的孔轴向连接，这意味着底物蛋白应该被展开以允许转运到消化室。ClpA内部的腔在结构上与其他伴侣的“安芬森笼”相当，可能在底物的展开中起作用。通过使用我们的ClpA模型和ClpP的晶体结构（Wang等人，1997年，《细胞》91卷，447 - 456页）来识别单个复合物中两个分子呈现的特定侧视图，获得了对称性不匹配的几何描述。这种相互作用的特征是一对关键亚基，每种蛋白质各一个。一个小的转动（8.6° = 2π/42；相当于4埃的位移）会将关键相互作用转移到另一对亚基。我们提出核苷酸水解导致旋转，促进底物蛋白的连续消化。
