Chapter 2: Determination of Reaction Intermediate Structures in Heme Proteins
Developments in structural biology and molecular biology have allowed increasingly detailed investigations of structure–function relationships. Although atomic-resolution structures of proteins are becoming more common, a growing number of structural studies have focused on the role played by dynamics and have sought to determine the structure of intermediates in protein reactions. These experiments have revealed the first atomic-level pictures of enzyme catalysis and the conformational motions required for biological function. This chapter uses the cryotrapping of reaction intermediates in horse heart myoglobin (Mb) to illustrate the methods utilized in determining the structures of reaction intermediates in protein systems. The techniques described here are applicable to a wide variety of heme proteins including Mb, hemoglobin, photosynthetic reaction centers, and cytochrome p450cam.