Correspondence to C. Andersen, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum) der Universität Würzburg, Am Hubland, D-97074 Würzburg, Germany
The porin RafY encoded by the raffinose plasmid pRSD2 of Escherichia coli forms a general diffusion pore and not a carbohydrate-specific porin
Article first published online: 25 DEC 2001
European Journal of Biochemistry
Volume 254, Issue 3, pages 679–684, June (II) 1998
How to Cite
Andersen, C., Krones, D., Ulmke, C., Schmid, K. and Benz, R. (1998), The porin RafY encoded by the raffinose plasmid pRSD2 of Escherichia coli forms a general diffusion pore and not a carbohydrate-specific porin. European Journal of Biochemistry, 254: 679–684. doi: 10.1046/j.1432-1327.1998.2540679.x
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Abbreviations. P(G), probability of occurrence of the conductance step with the single-channel conductance G; LDAO, lauryldimethylamine oxide.
- Issue published online: 25 DEC 2001
- Article first published online: 25 DEC 2001
- (Received 21 January/9 April 1998)
- Cited By
- raffinose transport;
- carbohydrate specificity;
- RafY channel;
- enteric bacteria;
- lipid bilayer membrane.
The gene rafY from the plasmid pRSD2, which enables Escherichia coli to grow on raffinose, was transferred into expression plasmid pUSL77. The protein was expressed in the porin-deficient Escherichia coli strain KS26 and was isolated and purified to homogeneity. The pure protein was reconstituted into lipid bilayer membranes. It formed an ion-permeable channel with a single-channel conductance of 2.9 nS of the open state in 1 M KCl, which is approximately twice of that of the general diffusion pores OmpF and OmpC of E. coli outer membrane. At lower pH the channel exhibited rapid flickering between three substates of the open channel. The RafY channel appears to be wide and water filled and has a small selectivity for cations over anions. Although RafY is part of an uptake and fermentation system for raffinose it does not contain a binding site for carbohydrates. Our results suggest that RafY is a general diffusion pore with a diameter, larger than that of the general diffusion porins OmpF and OmpC, that allows the diffusion of high-molecular-mass carbohydrates through the outer membrane.