Formaldehyde and photoactivatable cross-linking of the periplasmic binding protein to a membrane component of the histidine transport system of Salmonella typhimurium.
The histidine permease of Salmonella typhimurium consists of four protein components, one located in the periplasm and three in the cytoplasmic membrane. Genetic evidence indicated that the periplasmic protein interacts with the membrane proteins during transport. We have utilized two different methods to demonstrate that the periplasmic protein cross-links specifically to one of the membrane components, the Q protein. Formaldehyde, a water-soluble permeant molecule was used in vivo. Sulfosuccinimidyl 6-(4'-azido-2'-nitrophenylamino)hexanoate, a photoactivatable cross-linking reagent, was used in vitro in a reconstituted membrane vesicle system. Furthermore, we show that a mutant periplasmic protein, capable of binding substrate but not transporting it, is defective in cross-linking to the membrane protein, indicating this interaction to be a crucial step in the mechanism of transport.
Prossnitz
E
E
Department of Biochemistry, University of California, Berkeley.
Nikaido
K
K
Ulbrich
S J
SJ
Ames
G F
GF
D018528:ATP-Binding Cassette Transporters; D026923:Amino Acid Transport Systems, Basic; D001386:Azides / Q000378:metabolism*; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents / Q000378:metabolism*; D005557:Formaldehyde / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D010777:Photochemistry; D012486:Salmonella typhimurium / Q000201:enzymology*
D018528:ATP-Binding Cassette Transporters; D026923:Amino Acid Transport Systems, Basic; D001386:Azides; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D005557:Formaldehyde; C037733:N-succinimidyl-6-(4'-azido-2'-nitrophenylamino)hexanoate; C043828:histidine permease, Bacteria