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Signalling substitutions in the periplasmic domain of chemoreceptor Trg induce or reduce helical sliding in the transmembrane domain
Author(s) -
Beel Bryan D.,
Hazelbauer Gerald L.
Publication year - 2001
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2001.02446.x
Subject(s) - transmembrane domain , periplasmic space , transmembrane protein , biology , ligand (biochemistry) , biophysics , protein kinase domain , cysteine , biochemistry , amino acid , receptor , mutant , enzyme , escherichia coli , gene
We used in vivo oxidative cross‐linking of engineered cysteine pairs to assess conformational changes in the four‐helix transmembrane domain of chemoreceptor Trg. Extending previous work, we searched for and found a fourth cross‐linking pair that spanned the intrasubunit interface between transmembrane helix 1 (TM1) and its partner TM2. We determined the effects of ligand occupancy on cross‐linking rate constants for all four TM1–TM2 diagnostic pairs in conditions that allowed the formation of receptor–kinase complexes for the entire cellular complement of Trg. Occupancy altered all four rates in a pattern that implicated sliding of TM2 relative to TM1 towards the cytoplasm as the transmembrane signalling movement in receptor–kinase complexes. Transmembrane signalling can be reduced or induced by single amino acid substitutions in the ligand‐binding region of the periplasmic domain of Trg. We determined the effects of these substitutions on conformation in the transmembrane domain and on ligand‐induced changes using the diagnostic TM1–TM2 cysteine pairs. Effects on rates of in vivo cross‐linking showed that induced signalling substitutions altered the relative positions of TM1 and TM2 in the same way as ligand binding, and reduced signalling substitutions blocked or attenuated the ligand‐induced shift. These results provide strong support for the helical sliding model of transmembrane signalling.