title: High-resolution cryo-EM structure of the proteasome in complex with ADP-AlFx.
abstract: The 26S proteasome is an ATP-dependent dynamic 2.5 MDa protease that regulates numerous essential cellular functions through degradation of ubiquitinated substrates. Here we present a near-atomic-resolution cryo-EM map of the S. cerevisiae 26S proteasome in complex with ADP-AlFx. Our biochemical and structural data reveal that the proteasome-ADP-AlFx is in an activated state, displaying a distinct conformational configuration especially in the AAA-ATPase motor region. Noteworthy, this map demonstrates an asymmetric nucleotide binding pattern with four consecutive AAA-ATPase subunits bound with nucleotide. The remaining two subunits, Rpt2 and Rpt6, with empty or only partially occupied nucleotide pocket exhibit pronounced conformational changes in the AAA-ATPase ring, which may represent a collective result of allosteric cooperativity of all the AAA-ATPase subunits responding to ATP hydrolysis. This collective motion of Rpt2 and Rpt6 results in an elevation of their pore loops, which could play an important role in substrate processing of proteasome. Our data also imply that the nucleotide occupancy pattern could be related to the activation status of the complex. Moreover, the HbYX tail insertion may not be sufficient to maintain the gate opening of 20S core particle. Our results provide new insights into the mechanisms of nucleotide-driven allosteric cooperativity of the complex and of the substrate processing by the proteasome.
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Zhanyu
authors.initials: Z
authors.lastname: Ding
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Zhenglin
authors.initials: Z
authors.lastname: Fu
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.
authors.forename: Cong
authors.initials: C
authors.lastname: Xu
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.
authors.forename: Yifan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Yanxing
authors.initials: Y
authors.lastname: Wang
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Junrui
authors.initials: J
authors.lastname: Li
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Liangliang
authors.initials: L
authors.lastname: Kong
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.
authors.forename: Jinhuan
authors.initials: J
authors.lastname: Chen
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Na
authors.initials: N
authors.lastname: Li
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Rongguang
authors.initials: R
authors.lastname: Zhang
authors.affiliation: National Center for Protein Science Shanghai, State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences; University of Chinese Academy of Sciences, 333 Haike Road, Shanghai 201203, China.|Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Cong
mesh_terms: D000244:Adenosine Diphosphate / Q000737:chemistry* / Q000378:metabolism; D020285:Cryoelectron Microscopy; D008958:Models, Molecular; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000648:ultrastructure*
chemical_list: D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins; D000244:Adenosine Diphosphate; D046988:Proteasome Endopeptidase Complex