Traditional Biomolecular Structure Determination by NMR Spectroscopy Allows for Major Errors

Abstract

One of the major goals of structural genomics projects is to determine the three-dimensional structure of representative members of as many different fold families as possible. Comparative modeling is expected to fill the remaining gaps by providing structural models of homologs of the experimentally determined proteins. However, for such an approach to be successful it is essential that the quality of the experimentally determined structures is adequate. In an attempt to build a homology model for the protein dynein light chain 2A (DLC2A) we found two potential templates, both experimentally determined nuclear magnetic resonance (NMR) structures originating from structural genomics efforts. Despite their high sequence identity (96%), the folds of the two structures are markedly different. This urged us to perform in-depth analyses of both structure ensembles and the deposited experimental data, the results of which clearly identify one of the two models as largely incorrect. Next, we analyzed the quality of a large set of recent NMR-derived structure ensembles originating from both structural genomics projects and individual structure determination groups. Unfortunately, a visual inspection of structures exhibiting lower quality scores than DLC2A reveals that the seriously flawed DLC2A structure is not an isolated incident. Overall, our results illustrate that the quality of NMR structures cannot be reliably evaluated using only traditional experimental input data and overall quality indicators as a reference and clearly demonstrate the urgent need for a tight integration of more sophisticated structure validation tools in NMR structure determination projects. In contrast to common methodologies where structures are typically evaluated as a whole, such tools should preferentially operate on a per-residue basis. Three-dimensional biomolecular structures provide an invaluable source of biologically relevant information. To be able to learn the most of the wealth of information that these structures can provide us, it is of great importance that the quality and accuracy of the protein structure models deposited in the Protein Data Bank are as high as possible. In this work, the authors describe an analysis that illustrates that this is unfortunately not the case for many protein structures solved using nuclear magnetic resonance spectroscopy. They present an example in which two strikingly different models describing the same protein are analyzed using commonly available structure validation tools, and the results of this analysis show one of the two models to be incorrect. Subsequently, using a large set of recently determined structures, the authors demonstrate that unfortunately this example does not stand on its own. The analyses and examples clearly illustrate that relying solely on the experimental data to evaluate structural quality can provide a false sense of correctness and the combination of multiple sophisticated structure validation tools is required to detect the presence of errors in protein nuclear magnetic resonance structures.