title: A diffraction-quality protein crystal processed as an autophagic cargo.
abstract: Crystallization of proteins may occur in the cytosol of a living cell, but how a cell responds to intracellular protein crystallization remains unknown. We developed a variant of coral fluorescent protein that forms diffraction-quality crystals within mammalian cells. This expression system allowed the direct determination of its crystal structure at 2.9 Å, as well as observation of the crystallization process and cellular responses. The micron-sized crystal, which emerged rapidly, was a pure assembly of properly folded β-barrels and was recognized as an autophagic cargo that was transferred to lysosomes via a process involving p62 and LC3. Several lines of evidence indicated that autophagy was not required for crystal nucleation or growth. These findings demonstrate that in vivo protein crystals can provide an experimental model to study chemical catalysis. This knowledge may be beneficial for structural biology studies on normal and disease-related protein aggregation.
authors.affiliation: Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Wako, Saitama 351-0198, Japan; Department of Material Science, Japan Advanced Institute of Science and Technology, Nomi, Ishikawa 923-1292, Japan; PRESTO, Japan Science and Technology Agency (JST), 4-1-8 Hon-cho, Kawaguchi, Saitama 351-0198, Japan.
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Tsutsui
authors.affiliation: Laboratory for Integrative Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Yuka
authors.initials: Y
authors.lastname: Jinno
authors.affiliation: Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Wako, Saitama 351-0198, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Shoda
authors.affiliation: Laboratory for Integrative Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Akiko
authors.initials: A
authors.lastname: Tomita
authors.affiliation: Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Matsuda
authors.affiliation: Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Eiki
authors.initials: E
authors.lastname: Yamashita
authors.affiliation: Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Wako, Saitama 351-0198, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Katayama
authors.affiliation: Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nakagawa
authors.affiliation: Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Wako, Saitama 351-0198, Japan; Biotechnological Optic Research Team, RIKEN Center for Advanced Photonics, Wako, Saitama 351-0198, Japan. Electronic address: matsushi@brain.riken.jp.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Miyawaki
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D000818:Animals; D037421:Anthozoa / Q000737:chemistry*; D001343:Autophagy; D003460:Crystallization; D018360:Crystallography, X-Ray; D003600:Cytosol / Q000378:metabolism* / Q000648:ultrastructure; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009474:Neurons / Q000378:metabolism / Q000648:ultrastructure; D061251:Primary Cell Culture; D017510:Protein Folding; D017433:Protein Structure, Secondary; D021381:Protein Transport; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000071456:Sequestosome-1 Protein; D014961:X-Ray Diffraction
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D011994:Recombinant Proteins; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D049452:Green Fluorescent Proteins