Structural Mass Spectrometry of Proteins Using Hydroxyl Radical Based Protein Footprinting

摘要

Structural MS is a rapidly growing field with many applications in basic research and pharmaceutical drug development. In this feature article the overall technology is described and several examples of how hydroxyl radical based footprinting MS can be used to map interfaces, evaluate protein structure, and identify ligand dependent conformational changes in proteins are described. Driven by noncovalent interactions, polypeptide chains fold into a myriad of shapes that correspond to functional domains of biological significance. Therefore, assessment of protein structure is critical for understanding the functions of proteins at a molecular level. By comparing protein amino acid sequences and structures, scientists can classify proteins into family groups and deduce potential structure--function relationships by homology with proteins of known structure. Techniques such as X-ray crystallography and NMR spectroscopy can determine protein structures at atomic-level resolution. These experimentally determined structures of proteins, nucleic acids, and complex assemblies are collected into a database called the Protein Data Bank (PDB). Technical limitations of atomic resolution techniques have limited the direct determination of protein 3D structure when compared to the number of protein sequences available. Swiss-Prot, a protein sequence database, currently contains 500 000 unique sequences, whereas only 51 535 protein structures were registered in PDB as of 2009 and many of these PDB structures are highly redundant. Thus, experimental structures for the majority of known proteins do not exist. To fill this gap, computational modeling strategies based on homology and de novo prediction methods are necessary to provide structure--function predictions for systems in which the structure is poorly characterized.