Gf. Lee et al., DEDUCING THE ORGANIZATION OF A TRANSMEMBRANE DOMAIN BY DISULFIDE CROSS-LINKING - THE BACTERIAL CHEMORECEPTOR TRG, The Journal of biological chemistry, 269(47), 1994, pp. 29920-29927
The transmembrane domain of chemoreceptor Trg from Escherichia coli co
ntains four segments, two from each subunit of the homodimer. me used
site specific mutagenesis to introduce cysteines into those segments a
nd oxidative cross-linking of cysteine pairs to identify residues that
are near each other in space. Propensity for cross-linking was determ
ined for pairs of homologously placed cysteines in the two subunits of
the dimer at all 54 possible positions. Also, combinations of cystein
es were identified that readily oxidized to join heterologous segments
within or between monomers. These patterns of cross-licking were used
to develop a model for the three-dimensional structure of the transme
mbrane domain in which the four transmembrane segments are helices ass
ociated in a bundle, with stronger interactions near the periplasm and
weaker interactions near the cytoplasm. The striking similarity of th
is model to a model for the transmembrane domain of chemoreceptor Tar,
derived using the same experimental strategy, strengthens the notion
that a combination of comprehensive cysteine substitutions and analysi
s of patterns of disulfide cross-licking is sufficient to deduce a det
ailed three-dimensional structure for a transmembrane domain.