The meaning of alignment: lessons from structural diversity

We have taken alternative protein crystal structures and generated simulation snapshots to explicitly investigate the impact of structural changes on the alignments. We show that structural diversity has a significant effect on structural alignment. Moreover, we observe alignment inconsistencies even for modest spatial divergence, implying that the biological interpretation of alignments is less straightforward than commonly assumed. A salient example is the GroES 'mobile loop' where sub-？ngstrom variations give rise to contradictory sequence alignments.A comprehensive treatment of ambiguous alignment regions is crucial for further development of structural alignment applications and for the representation of alignments in general. For this purpose we have developed an on-line database containing our data and new ways of visualizing alignment inconsistencies, which can be found at http://www.ibi.vu.nl/databases/stralivari webcite.Sequence comparison has become a major tool for biological research in the post-genomic era, forming the basis for functional annotation, classification, and analysis of evolutionary relationships. At the residue level, however, the relation between sequence, structure and function can often be obscure, and examples abound of proteins with a clear functional and homologous relationship but sharing negligible similarity at the sequence level.Structural alignment therefore is the method of choice for reliable homology assessment and derived features like functional classification and phylogeny. This importance is reflected in the number of tools available for structural alignment, such as DALI [1], SSAP [2], STRUCTAL [3], MAMMOTH [4], CE [5] and COMPARER [6] (for recent reviews on the topic, see Kolodny et al. [7] and Mayr et al. [8]). Databases for functional classification such as CATH [9], FSSP [10] and PASS2 [11] each derive directly from the use of one or more of these methods, whereas for SCOP expert input in the structural classificati