Journal of Bacteriology, November 2003, p. 6695-6701, Vol. 185, No. 22
0021-9193/03/$08.00+0 DOI: 10.1128/JB.185.22.6695-6701.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
Structural and Topographical Studies of the Type IV Bundle-Forming Pilus Assembly Complex of Enteropathogenic Escherichia coli
Jaiweon Hwang, David Bieber,
Sandra W. Ramer,
Cheng-Yen Wu, and Gary K. Schoolnik*
Departments of Medicine (Infectious Diseases and Geographic Medicine) and Microbiology & Immunology, Stanford Medical School, Stanford, California 94305
Received 29 May 2003/
Accepted 21 August 2003
The type IV bundle-forming pili (BFP) of enteropathogenic Escherichia coli (EPEC) are required for virulence in orally challenged human volunteers and for the localized adherence and autoaggregation in vitro phenotypes. BFP filament biogenesis and function are encoded by the 14-gene bfp operon. The BFP assembly complex, containing a BfpB-His6 fusion protein, was chemically cross-linked in situ, and the complex was then purified from BFP-expressing EPEC by a combination of nickel- and BfpB antibody-based affinity chromatography. Characterization of the isolated complex by immunoblotting using BFP protein-specific antibodies showed that at least 10 of the 14 proteins specified by the bfp operon physically interact to form an oligomeric complex. Proteins localized to the outer membrane, inner membrane, and periplasm are within this complex, thus demonstrating that the complex spans the periplasmic space. A combination of immunofluorescence and immuno-gold thin-section transmission electron microscopy studies localized this complex to one pole of the cell.
* Corresponding author. Mailing address: Departments of Medicine (Infectious Diseases and Geographic Medicine) and Microbiology & Immunology, Beckman Center, Rm. 241, Stanford Medical School, Stanford, CA 94305. Phone: (650) 723-8158. Fax: (650) 723-1399. E-mail: schoolni{at}cmgm.stanford.edu.
Present address: San Jose State University, San Jose, CA 95192.
Present address: Genencor International, Inc., Palo Alto, CA 94304.
Journal of Bacteriology, November 2003, p. 6695-6701, Vol. 185, No. 22
0021-9193/03/$08.00+0 DOI: 10.1128/JB.185.22.6695-6701.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
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Copyright © 2003 by the American Society for Microbiology. All rights reserved.