Thermodynamic characterization of OsGID1‐gibberellin binding using calorimetry and docking simulations

Gibberellins (GAs) are phytohormones regulating various developmental processes in plants. In rice, the initial GA‐signaling events involve the binding of a GA to the soluble GA receptor protein, GID1. Although X‐ray structures for certain GID1/GA complexes have recently been determined, an examination of the complexes does not fully clarify how GID1s discriminate among different GAs. Herein, we present a study aimed at defining the types of forces important to binding via a combination of isothermal titration calorimetry (ITC) and computational docking studies that employed rice GID1 (OsGID1), OsGID1 mutants, which were designed to have a decreased possible number of hydrogen bonds with bound GA, and GA variants. We find that, in general, GA binding is enthalpically driven and that a hydrogen bond between the phenolic hydroxyl of OsGID1 Tyr134 and the C‐3 hydroxyl of a GA is a defining structural element. A hydrogen‐bond network that involves the C‐6 carboxyl of a GA that directly hydrogen bonds the hydroxyl of Ser198 and indirectly, via a two‐water‐molecule network, the phenolic hydroxyl of Tyr329 and the NH of the amide side‐chain of Asn255 is also important for GA binding. The binding of OsGID1 by GA 1 is the most enthalpically driven association found for the biologically active GAs evaluated in this study. This observation might be a consequence of a hydrogen bond formed between the hydroxyl at the C‐13 position of GA 1 and the main chain carbonyl of OsGID1 Phe245. Our results demonstrate that by combining ITC experiments and computational methods much can be learned about the thermodynamics of ligand/protein binding. Copyright © 2010 John Wiley & Sons, Ltd.