This Article 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 Thomas, W D Articles by Welch, R A Search for Related Content PubMed PubMed Citation Articles by Thomas, W D, Jr Articles by Welch, R A

research-article

A heterologous membrane protein domain fused to the C-terminal ATP-binding domain of HlyB can export Escherichia coli hemolysin.

Department of Medical Microbiology and Immunology, University of Wisconsin-Madison 53706.

ABSTRACT

The hydrophobic-rich NH2-terminal 34 amino acids of a tetracycline resistance determinant (TetC) were fused to the COOH-terminal 240 amino acids of the hemolysin transporter, HlyB, which contains a putative ATP-binding domain. This hybrid protein replaced the NH2-terminal 467-amino-acid portion of HlyB and could still export the Escherichia coli hemolysin (HlyA). Export by the hybrid protein was approximately 10% as efficient as transport by HlyB. Extracellular secretion of HlyA by the TetC-HlyB hybrid required HlyD and TolC. The extracellular and periplasmic levels of beta-galactosidase and beta-lactamase in strains that produced the hybrid were similar to the levels in controls. Thus, HlyA transport was specific and did not appear to be due to leakage of cytoplasmic contents alone. Antibodies raised against the COOH terminus of HlyB reacted with the hybrid protein, as well as HlyB. HlyB was associated with membrane fractions, while the hybrid protein was found mainly in soluble extracts. Cellular fractionation studies were performed to determine whether transport by the hybrid occurred simultaneously across both membranes like wild-type HlyA secretion. However, we found that HlyA was present in the periplasm of strains that expressed the TetC-HlyB hybrid. HlyA remained in the periplasm unless the hlyD and tolC gene products were present in addition to the hybrid.

This article has been cited by other articles:

• Leduc, I., White, C. D., Nepluev, I., Throm, R. E., Spinola, S. M., Elkins, C. (2008). Outer Membrane Protein DsrA Is the Major Fibronectin-Binding Determinant of Haemophilus ducreyi. Infect. Immun. 76: 1608-1616 [Abstract] [Full Text]  
• Palacios, J. L., Zaror, I., Martínez, P., Uribe, F., Opazo, P., Socías, T., Gidekel, M., Venegas, A. (2001). Subset of Hybrid Eukaryotic Proteins Is Exported by the Type I Secretion System of Erwinia chrysanthemi. J. Bacteriol. 183: 1346-1358 [Abstract] [Full Text]  
• Moxley, R. A., Berberov, E. M., Francis, D. H., Xing, J., Moayeri, M., Welch, R. A., Baker, D. R., Barletta, R. G. (1998). Pathogenicity of an Enterotoxigenic Escherichia coli Hemolysin (hlyA) Mutant in Gnotobiotic Piglets. Infect. Immun. 66: 5031-5035 [Abstract] [Full Text]