This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Mori, H. Articles by Ito, K. Search for Related Content PubMed PubMed Citation Articles by Mori, H. Articles by Ito, K.

 Previous Article  |  Next Article 

Journal of Bacteriology, June 2004, p. 3960-3969, Vol. 186, No. 12
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.12.3960-3969.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Mutational Analysis of Transmembrane Regions 3 and 4 of SecY, a Central Component of Protein Translocase

Hiroyuki Mori, Naomi Shimokawa, Yasunari Satoh, and Koreaki Ito^*

Institute for Virus Research, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan

Received 9 December 2003/ Accepted 16 March 2004

The SecYEG heterotrimeric membrane protein complex functions as a channel for protein translocation across the Escherichia coli cytoplasmic membrane. SecY is the central subunit of the SecYEG complex and contains 10 transmembrane segments (TM1 to TM10). Previous mutation studies suggested that TM3 and TM4 are particularly important for SecY function. To further characterize TM3 and TM4, we introduced a series of cysteine-scanning mutations into these segments. With one exception (an unstable product), all the mutant proteins complemented the cold-sensitive growth defect of the secY39 mutant. A combination of this secY mutation and the secG deletion resulted in synthetic lethality, and the TM3 and TM4 SecY cysteine substitution mutations were examined for their ability to complement this lethality. Although they were all positive for complementation, some of the complemented cells exhibited significant retardation of protein export. The substitution-sensitive residues in TM3 can be aligned to one side of the alpha-helix, and those in TM4 revealed a tendency for residues closer to the cytosolic side of the membrane to be more severely affected. Disulfide cross-linking experiments identified a specific contact point for TM3 and SecG TM2 as well as for TM4 and SecG TM1. Thus, although TM3 and TM4 do not contain any single residue that is absolutely required, they include functionally important helix surfaces and specific contact points with SecG. These results are discussed in light of the structural information available for the SecY complex.

---

* Corresponding author. Mailing address: Institute for Virus Research, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan. Phone: (75) 751-4015. Fax: (75) 771-5699. E-mail: kito{at}virus.kyoto-u.ac.jp.

---

Journal of Bacteriology, June 2004, p. 3960-3969, Vol. 186, No. 12
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.12.3960-3969.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Nagler, C., Nagler, G., Kuhn, A. (2007). Cysteine Residues in the Transmembrane Regions of M13 Procoat Protein Suggest that Oligomeric Coat Proteins Assemble onto Phage Progeny. J. Bacteriol. 189: 2897-2905 [Abstract] [Full Text]  
• Shimohata, N., Nagamori, S., Akiyama, Y., Kaback, H. R., Ito, K. (2007). SecY alterations that impair membrane protein folding and generate a membrane stress. JCB 176: 307-317 [Abstract] [Full Text]  
• Mori, H., Ito, K. (2006). Different modes of SecY-SecA interactions revealed by site-directed in vivo photo-cross-linking. Proc. Natl. Acad. Sci. USA 103: 16159-16164 [Abstract] [Full Text]