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Transport of Vitamin B₁₂ in Escherichia coli: Common Receptor Sites for Vitamin B₁₂ and the E Colicins on the Outer Membrane of the Cell Envelope

¹ Departments of Biochemistry and Microbiology, University of Virginia School of Medicine, Charlottesville, Virginia 22901

ABSTRACT

The first step in the transport of cyanocobalamin (CN-B₁₂) by cells of Escherichia coli was shown previously to consist of binding of the B₁₂ to specific receptor sites located on the outer membrane of the cell envelope. In this paper, evidence is presented that these B₁₂ receptor sites also function as the receptors for the E colicins, and that there is competition between B₁₂ and the E colicins for occupancy of these sites. The cell strains used were E. coli KBT001, a methionine/B₁₂ auxotroph, and B₁₂ transport mutants derived from strain KBT001. Colicins E1 and E3 inhibited binding of B₁₂ to the outer membrane B₁₂ receptor sites, and CN-B₁₂ protected cells against these colicins. Half-maximal protection was given by CN-B₁₂ concentrations in the range of 1 to 6 nM, depending upon the colicin concentration used. Colicin E1 competitively inhibited the binding of ⁵⁷Co-labeled CN-B₁₂ to isolated outer membrane particles. Functional colicin E receptor sites were found in cell envelopes from cells of only those strains that possessed intact B₁₂ receptors. Colicin K did not inhibit the binding of B₁₂ to the outer membrane receptor sites, and no evidence was found for any identity between the B₁₂ and colicin K receptors. However, both colicin K and colicin E1 inhibited the secondary phase of B₁₂ transport, which is believed to consist of the energy-coupled movement of B₁₂ across the inner membrane.

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