Contributions of Conformational Flexibility to High-Affinity Zinc Binding in the Solute Binding Protein AztC.

Bacteria rely on ATP binding cassette (ABC) transporters for the import of various nutrients. Bacterial ABC importers utilize an extracellular solute binding protein (SBP) to bind the substrate with high affinity and specificity and deliver it to the membrane permease for transport. The essential metals iron, manganese, and zinc are bound and transported by the cluster A-I SBPs. Crystal structures exist for the metal-bound and metal-free forms of several cluster A-I SBPs that show relatively subtle conformational changes that accompany metal binding. Recent solution studies and molecular dynamics simulations indicate a more complex conformational landscape for the cluster A-I SBPs, suggesting that changes in protein dynamics upon metal binding may have an important role in recognition by the membrane permease and effective transport. Here, we investigate conformational states and dynamics in the cluster A-I SBP AztC from, characterizing its unusual intrinsic fluorescence behavior and thermodynamics of zinc binding. These data suggest a dynamic equilibrium of at least two conformational states in the apo form and compensatory changes in the holo that provide for a significant entropic contribution to zinc binding. Correlation with available crystal structures suggests that the formation of a Trp-Phe π-stacking interaction in the metal-bound form may mediate the observed changes in fluorescence. The conformational dynamics identified here for AztC are likely applicable to other cluster A-I SBPs with relevance to their exploitation as potential antibiotic drug targets.