Transport of vitamin B12 in Escherichia coli. Some observations on the roles of the gene products of BtuC and TonB.

The transport of vitamin B I Z (cyanocobalamin, CN-Cbl) in Escherichia coli requires the proton motive force and functional products from the genes BtuB, TonB, and BtuC.E. coli cells that contain the btuC mutation are defective in CN-Cbl transport across the inner membrane and accumulate large concentrations of CN-Cbl in the periplasmic space.This uptake of CN-Cbl into the periplasmic space requires the TonB gene product, the proton motive force, and two functional sites on the outer membrane cobalamin (Cbl) receptor protein (the Cbl-binding site and a second site which is necessary for transfer of Cbl from the receptor towards the interior of the cell).We propose that the TonB gene product, the proton motive force, and the second domain on the receptor interact to increase the rate of dissociation of the receptor-Cbl complex, thus releasing Cbl into the periplasmic space.We also suggest that a reasonable mechanism for this catalyzed release of Cbl is via the interaction of the second functional domain on the receptor with a diffusible messenger that is generated in the periplasmic space by the combined action of the proton motive force and the inner membrane TonB protein.Arsenate was found to have two distinct effects upon CN-Cbl uptake, a 3-to 12-fold stimulation of CN-Cbl transport across the outer membrane, which was seen in btuC cells, and inhibition of inner membrane transport of CN-Cbl in wild type cells.The latter effect resulted in the periplasmic accumulation of CN-Cbl and may indicate a requirement for phosphate bond energy in Cbl transport across the inner membrane.A variety of other reagents, including arsenite, diamide, N-ethylmaleimide, and sucrose, gave small stimulations (usually <2-fold) of CN-Cbl uptake in btuC cells, possibly as a result of increasing the size of the periplasmic space.Most of the genes involved in the transport of cyanocobalamin (CN-Cbl) across the cell envelope of Escherichia coli are shared with other membrane processes.The first step in this transport process is the binding of Cbl' to the outer membrane Cbl receptor, the protein component of which is the product of the BtuB gene and also functions as the receptor for bacteriophage BF23 and the E colicins (1,2).