A binding mechanism in protein–nucleotide interactions: Implication for U1A RNA binding

Abstract

We present a close electronic view of the protein-base interface for the N-terminal domain of the human protein U1A. Combining accurate mixed quantum mechanics͞molecular mechanics techniques and protein structure prediction methods, we provide a detailed electronic structure description of the protein-RNA stacking interactions. Our analysis indicates the evolution of the protein structure optimizing the interaction between Asp-92 and the RNA bases. The results show a direct coupling of the C-terminal tail and Asp-92, providing a direct rationalization of the experimentally determined role of the C-terminal domain in RNA binding. Here, we propose a mechanism where a protein side chain, with a delocalized electronic pi system, assists in the nucleotide binding. The binding mechanism involves a short-range interaction of the side chain with the nucleotide base and an electronic long-range interaction through a sandwich-stacking motif. The structural motif of the binding mechanism is observed in similar protein-RNA interactions and in various protein-ATP-binding sites.ATP binding ͉ quantum mechanics͞molecular mechanics ͉ stacking ͉ aromatic T he mechanisms of protein-nucleotide association constitute a central intermolecular interaction. Such mechanisms, for example, regulate DNA, RNA, and NTP recognition and binding. As pointed out recently by Williamson (1), there is a large complexity of induced fit processes, requiring conformational change in the proteins, in the ligands, or in both. These dynamical processes, responsible for the induced affinity and specificity, are far from being understood. A common feature in ssDNA, RNA, and NTP binding is the direct interaction of the protein with the nucleotides' bases. Recent structural analyses have underlined the main features involved in these protein-base interactions (2, 3). These studies revealed the importance of van der Waal interactions, aromatic stacking interactions in particular, in the stabilization of the protein-base interaction.The ribonucleoprotein domain, also known as the RNAbinding domain or the RNA-recognition motif, binds RNA by using extensive interactions through its characteristic fourstranded ␤-sheet structure (Fig. 1). Among the members of this motif, the N-terminal domain of the human protein U1A is the most studied (4-29). The isolated domain binds to a short RNA hairpin with high affinity and specificity. The importance of stacking interactions in RNA binding is confirmed by the aromatic nature of three of the most highly conserved residues in the RNA-recognition motif (30). Baranger and coworkers (11) recently obtained 5.5 kcal͞mol destabilization of the protein-RNA complex when mutating Phe-56 to alanine. Subsequent studies by the same group indicated that stacking (hydrophobic) interactions are able to compensate for the loss of hydrogenbonding-capable functional groups in the RNA bases (5). Recent kinetic data by Laird-Offringa and coworkers (6) supported a rapid initial association based on electrostatic interactions, followed by a subs...

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