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

 Previous Article  |  Next Article 

Journal of Bacteriology, December 2007, p. 8961-8972, Vol. 189, No. 24
0021-9193/07/$08.00+0     doi:10.1128/JB.01365-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Isolation of Cold-Sensitive yidC Mutants Provides Insights into the Substrate Profile of the YidC Insertase and the Importance of Transmembrane 3 in YidC Function{triangledown}

Jijun Yuan,1 Gregory J. Phillips,2 and Ross E. Dalbey1*

Department of Chemistry, The Ohio State University, Columbus, Ohio 43210,1 Department of Veterinary Microbiology, Iowa State University, Ames, Iowa 500112

Received 21 August 2007/ Accepted 28 September 2007

YidC, a 60-kDa integral membrane protein, plays an important role in membrane protein insertion in bacteria. YidC can function together with the SecYEG machinery or operate independently as a membrane protein insertase. In this paper, we describe two new yidC mutants that lead to a cold-sensitive phenotype in bacterial cell growth. Both alleles impart a cold-sensitive phenotype and result from point mutations localized to the third transmembrane (TM3) segment of YidC, indicating that this region is crucial for YidC function. We found that the yidC(C423R) mutant confers a weak phenotype on membrane protein insertion while a yidC(P431L) mutant leads to a stronger phenotype. In both cases, the affected substrates include the Pf3 coat protein and ATP synthase F1Fo subunit c (FoC), while CyoA (the quinol binding subunit of the cytochrome bo3 quinol oxidase complex) and wild-type procoat are slightly affected or not affected in either cold-sensitive mutant. To determine if the different substrates require various levels of YidC activity for membrane insertion, we performed studies where YidC was depleted using an arabinose-dependent expression system. We found that –3M-PC-Lep (a construct with three negatively charged residues inserted into the middle of the procoat-Lep [PC-Lep] protein) and Pf3 P2 (a construct with the Lep P2 domain added at the C terminus of Pf3 coat) required the highest amount of YidC and that CyoA-N-P2 (a construct with the amino-terminal part of CyoA fused to the Lep P2 soluble domain) and PC-Lep required the least, while FoC required moderate YidC levels. Although the cold-sensitive mutations can preferentially affect one substrate over another, our results indicate that different substrates require different levels of YidC activity for membrane insertion. Finally, we obtained several intragenic suppressors that overcame the cold sensitivity of the C423R mutation. One pair of mutations suggests an interaction between TM2 and TM3 of YidC. The studies reveal the critical regions of the YidC protein and provide insight into the substrate profile of the YidC insertase.

* Corresponding author. Mailing address: Department of Chemistry, The Ohio State University, 100 W. 18th Ave., Columbus, OH 43210. Phone: (614) 292-2384. Fax: 614-292-1532. E-mail: dalbey{at}chemistry.ohio-state.edu

{triangledown} Published ahead of print on 12 October 2007.

Journal of Bacteriology, December 2007, p. 8961-8972, Vol. 189, No. 24
0021-9193/07/$08.00+0     doi:10.1128/JB.01365-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Yu, Z., Koningstein, G., Pop, A., Luirink, J. (2008). The Conserved Third Transmembrane Segment of YidC Contacts Nascent Escherichia coli Inner Membrane Proteins. J. Biol. Chem. 283: 34635-34642 [Abstract] [Full Text]  
  • Ravaud, S., Stjepanovic, G., Wild, K., Sinning, I. (2008). The Crystal Structure of the Periplasmic Domain of the Escherichia coli Membrane Protein Insertase YidC Contains a Substrate Binding Cleft. J. Biol. Chem. 283: 9350-9358 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»