Abstract

Newly synthesized membrane proteins must be accurately inserted into the membrane, folded and assembled for proper functioning. The protein YidC inserts its substrates into the membrane, thereby facilitating membrane protein assembly in bacteria; the homologous proteins Oxa1 and Alb3 have the same function in mitochondria and chloroplasts, respectively1,2. In the bacterial cytoplasmic membrane, YidC functions as an independent insertase and a membrane chaperone in cooperation with the translocon SecYEG3,4,5. Here we present the crystal structure of YidC from Bacillus halodurans, at 2.4 Å resolution. The structure reveals a novel fold, in which five conserved transmembrane helices form a positively charged hydrophilic groove that is open towards both the lipid bilayer and the cytoplasm but closed on the extracellular side. Structure-based in vivo analyses reveal that a conserved arginine residue in the groove is important for the insertion of membrane proteins by YidC. We propose an insertion mechanism for single-spanning membrane proteins, in which the hydrophilic environment generated by the groove recruits the extracellular regions of substrates into the low-dielectric environment of the membrane.

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Accessions

Primary accessions

Data deposits

The atomic coordinates and structure factors for YidC27–266 and YidC27–267 have been deposited in the Protein Data Bank under accession numbers 3WO6 and 3WO7, respectively.

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Acknowledgements

We wish to thank K. Watanabe from Shoko Scientific for assistance with the SEC-MALLS experiments; T. Nishizawa, T. Higuchi, H. E. Kato, M. Hattori, R. Ishii and H. Nishimasu for discussions; A. Kurabayashi, H. Nakamura, S. Hibino, T. Takino and C. Tsutsumi for technical support; A. Nakashima and R. Yamazaki for secretarial assistance; the RIKEN BioResource Center for providing B. halodurans genomic DNA; the RIKEN Integrated Cluster of Clusters (RICC) for providing computational resources; and the beamline staff members at BL32XU of SPring-8 for technical assistance during data collection. The synchrotron radiation experiments were performed at BL32XU of SPring-8 (proposal no. 2011A1125, 2011A1139, 2011B1062, 2011B1280, 2012A1093, 2012A1201, 2012B1146, 2012B1162 and 2013A1128), with approval from RIKEN. This work was supported by the Platform for Drug Discovery, Informatics and Structural Life Science by the Ministry of Education, Culture, Sports, Science and Technology (MEXT), by JSPS KAKENHI (grant no. 20247020, 20523517, 24687016, 24102503, 24121704, 24227004, 24657095, 25291006, 25291009 and 25660073), by the FIRST program, by PRESTO, by the JST, by a Grant-in-Aid for JSPS Fellows, by a grant for the HPCI STRATEGIC PROGRAM Computational Life Science and Application in Drug Discovery and Medical Development from MEXT, and by grants from the Private University Strategic Research Foundation Support Program (MEXT), the Nagase Science and Technology Foundation, and the Astellas Foundation for Research on Metabolic Disorders.

Author information

Author notes

    • Kaoru Kumazaki
    •  & Shinobu Chiba

    These authors contributed equally to this work.

Affiliations

  1. Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan

    • Kaoru Kumazaki
    • , Mizuki Takemoto
    • , Ryuichiro Ishitani
    •  & Osamu Nureki
  2. Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan

    • Kaoru Kumazaki
    • , Mizuki Takemoto
    • , Naoshi Dohmae
    • , Ryuichiro Ishitani
    •  & Osamu Nureki
  3. Faculty of Life Sciences, Kyoto Sangyo University, Motoyama, Kamigamo, Kita-ku, Kyoto 603-8555, Japan

    • Shinobu Chiba
    •  & Koreaki Ito
  4. Department of Systems Biology, Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama-cho, Ikoma, Nara 630-0192, Japan

    • Arata Furukawa
    • , Yasunori Sugano
    • , Yoshiki Tanaka
    •  & Tomoya Tsukazaki
  5. Cryobiofrontier Research Center, Faculty of Agriculture, Iwate University, 3-18-8 Ueda, Morioka, Iwate 020-8550, Japan

    • Ken-ichi Nishiyama
  6. Theoretical Molecular Science Laboratory, RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan

    • Takaharu Mori
    •  & Yuji Sugita
  7. SR Life Science Instrumentation Unit, RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan

    • Kunio Hirata
  8. Department of Cell Biology, Graduate School of Medicine, Kyoto University, Yoshidakonoe-cho, Sakyo-ku, Kyoto 606-8501, Japan

    • Yoshiko Nakada-Nakura
  9. Department of Bioengineering Sciences, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan

    • Andrés D. Maturana
  10. Institute for Virus Research, Kyoto University, Shogoin Kawara-cho, Sakyo-ku, Kyoto 606-8507, Japan

    • Hiroyuki Mori
  11. Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503, Japan

    • Fumio Arisaka
  12. JST, PRESTO, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan

    • Tomoya Tsukazaki

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Contributions

K.K. performed the crystallization and structure determination. S.C. performed the genetic analyses. K.K., A.F., K.-I.N., Y. Sugano, A.D.M., Y.T., H.M. and T.T. performed the functional analysis. M.T., T.M., Y. Sugita and R.I. performed the molecular dynamics simulation. K.K., N.D. and F.A. identified the molecular mass. K.H., Y.N.-N., R.I., T.T. and O.N. assisted with the structure determination. K.K., S.C., K.I., R.I., T.T. and O.N. wrote the manuscript. T.T. and O.N. directed and supervised all of the research.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Tomoya Tsukazaki or Osamu Nureki.

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https://doi.org/10.1038/nature13167

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