Prediction and experimental validation of enzyme substrate specificity in protein structures

Many proteins solved by Structural Genomics have low sequence identity to other proteins and cannot be assigned functions. To address this problem, we present a computational approach that creates structural motifs of a few evolutionarily important residues, and these motifs probe local geometric and evolutionary similarities in other protein structures to detect functional similarities. This approach does not require prior knowledge of functional mechanisms and is highly accurate in computational benchmarks when annotations rely on homologs with low sequence identity. We further demonstrate the accuracy of this approach using biochemical and mutagenesis studies to validate two predictions of unannotated proteins.