title: Transparent Window Vibrational Probes for the Characterization of Proteins With High Structural and Temporal Resolution.
abstract: Vibrational spectroscopy provides a direct route to the physicochemical characterization of molecules. While both IR and Raman spectroscopy have been used for decades to provide detailed characterizations of small molecules, similar studies with proteins are largely precluded due to spectral congestion. However, the vibrational spectra of proteins do include a "transparent window", between ∼1800 and ∼2500 cm-1, and progress is now being made to develop site-specifically incorporated carbon-deuterium (C-D), cyano (CN), thiocyanate (SCN), and azide (N3) "transparent window vibrational probes" that absorb within this window and report on their environment to facilitate the characterization of proteins with small molecule-like detail. This Review opens with a brief discussion of the advantages and limitations of conventional vibrational spectroscopy and then discusses the strengths and weaknesses of the different transparent window vibrational probes, methods by which they may be site-specifically incorporated into peptides and proteins, and the physicochemical properties they may be used to study, including electrostatics, stability and folding, hydrogen bonding, protonation, solvation, dynamics, and interactions with inhibitors. The use of the probes to vibrationally image proteins and other biomolecules within cells is also discussed. We then present four case studies, focused on ketosteroid isomerase, the SH3 domain, dihydrofolate reductase, and cytochrome c, where the transparent window vibrational probes have already been used to elucidate important aspects of protein structure and function. The Review concludes by highlighting the current challenges and future potential of using transparent window vibrational probes to understand the evolution and function of proteins and other biomolecules.
authors.affiliation: Department of Chemistry, The Scripps Research Institute , La Jolla, California 92037, United States.
authors.forename: Ramkrishna
authors.initials: R
authors.lastname: Adhikary
authors.affiliation: Department of Chemistry, The Scripps Research Institute , La Jolla, California 92037, United States.
authors.forename: Jörg
authors.initials: J
authors.lastname: Zimmermann
authors.affiliation: Department of Chemistry, The Scripps Research Institute , La Jolla, California 92037, United States.
authors.forename: Floyd E
authors.identifier: 0000-0001-6317-1315
authors.initials: FE
authors.lastname: Romesberg
mesh_terms: D015335:Molecular Probes / Q000737:chemistry*; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D013055:Spectrophotometry, Infrared; D013057:Spectrum Analysis / Q000379:methods; D014732:Vibration
chemical_list: D015335:Molecular Probes; D011506:Proteins