Detergents affect the solution-phase structural dynamics of LeuT and have an impact on the observed HDX rate. Addition of substrates leads to perturbed HDX, relative to the apo state, in several LeuT regions. Presented HDX data is consistent with existing structural models and pinpoints LeuT segments involved in substrate transport. HDX-MS enables the detection of slow and correlated unfolding motions in transmembrane helices. This helical unwinding might be important in the outward-to-inward transition of LeuT. Substrates markedly modulate the rate of unfolding. Individual transmembrane helices (TM1a/TM5i and TM6a/TM7i) in LeuT seem to be conformationally coupled.
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