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Biophys. J. BioFAST: First Published August 11, 2006. doi:10.1529/biophysj.106.088781
© 2006 by the Biophysical Society.

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CHANNELS, RECEPTORS, AND ELECTRICAL SIGNALING

"Pore Characteristics of Nontypeable Haemophilus influenzae Outer Membrane Protein P5 in Planar Lipid Bilayers"

Eleonora Zakharian 1 and Rosetta N Reusch 2*

1 University of Medicine and Dentistry of New Jersey
2 Michigan State University

* To whom correspondence should be addressed. E-mail: rnreusch{at}msu.edu.

Submitted on May 11, 2006
Revised on July 11, 2006
Accepted on 31 July 2006


  Abstract
The structure of outer membrane protein P5 of nontypeable (acapsulate) Haemophilus influenzae (NTHi P5), a homolog of Escherichia coli OmpA, was investigated by observing its pore characteristics in planar lipid bilayers. Recombinant NTHi P5 was overexpressed in E. coli and purified using ionic detergent, lithium dodecyl sulfate (LDS-P5), or nonionic detergent, octylglucoside (OG-P5). LDS-P5 and OG- P5 could not be distinguished by their migration on SDS-PAGE gels; however, when incorporated into planar bilayers of DPhPC between symmetric aqueous solutions of 1 M KCl at 22 °C, LDS- P5 formed narrow pores (58 ± 6 pS) with low open probability whereas OG-P5 formed large pores (1.1 ± 0.1 nS) with high open probability (0.99). LDS-P5 narrow pores were gradually and irreversibly transformed into large pores, indistinguishable from those formed by OG-P5, at temperatures ≥ 40 °C; the process took 4-6 hrs at 40 °C or 35-45 min at 42 °C. Large pores were stable to changes in temperatures; however, large pores were rapidly converted to narrow pores when exposed to LDS at room temperatures, indicating acute sensitivity of this conformer to ionic detergent. These studies suggest that narrow pores are partially denatured forms and support the premise that the native conformation of NTHi P5 is that of a large monomeric pore.

Key Words: H. influenzae, NTHi P5, OmpA homolog, planar lipid bilayers, pore, structure transition




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E. Zakharian and R. N. Reusch
Haemophilus influenzae Outer Membrane Protein P5 Is Associated with Inorganic Polyphosphate and Polyhydroxybutyrate
Biophys. J., January 15, 2007; 92(2): 588 - 593.
[Abstract] [Full Text] [PDF]


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Copyright © 2006 by the Biophysical Society.