title: Studying submicrosecond protein folding kinetics using a photolabile caging strategy and time-resolved photoacoustic calorimetry.
abstract: Kinetic measurement of protein folding is limited by the method used to trigger folding. Traditional methods, such as stopped flow, have a long mixing dead time and cannot be used to monitor fast folding processes. Here, we report a compound, 4-(bromomethyl)-6,7-dimethoxycoumarin, that can be used as a "photolabile cage" to study the early stages of protein folding. The folding process of a protein, RD1, including kinetics, enthalpy, and volume change, was studied by the combined use of a phototriggered caging strategy and time-resolved photoacoustic calorimetry. The cage caused unfolding of the photolabile protein, and then a pulse UV laser (∼10(-9) s) was used to break the cage, leaving the protein free to refold and allowing the resolving of two folding events on a nanosecond time scale. This strategy is especially good for monitoring fast folding proteins that cannot be studied by traditional methods.
authors.affiliation: Institute of Biological Chemistry, Academia Sinica, Taipei 11529, Taiwan.
authors.forename: Hsin-Liang
authors.initials: HL
authors.lastname: Chen
authors.forename: Jack C-C
authors.initials: JC
authors.lastname: Hsu
authors.forename: Man Hoang
authors.initials: MH
authors.lastname: Viet
authors.forename: Mai Suan
authors.initials: MS
authors.lastname: Li
authors.forename: Chin-Kun
authors.initials: CK
authors.lastname: Hu
authors.forename: Chia-Hsun
authors.initials: CH
authors.lastname: Liu
authors.forename: Frederick Y
authors.initials: FY
authors.lastname: Luh
authors.forename: Silvia S-W
authors.initials: SS
authors.lastname: Chen
authors.forename: Evan S-H
authors.initials: ES
authors.lastname: Chang
authors.forename: Andrew H-J
authors.initials: AH
authors.lastname: Wang
authors.forename: Min-Feng
authors.initials: MF
authors.lastname: Hsu
authors.forename: Wunshain
authors.initials: W
authors.lastname: Fann
authors.forename: Rita P-Y
authors.initials: RP
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D021343:Antifreeze Proteins, Type III / Q000737:chemistry*; D002151:Calorimetry; D002942:Circular Dichroism; D003198:Computer Simulation; D003374:Coumarins / Q000737:chemistry*; D007700:Kinetics; D007834:Lasers; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017510:Protein Folding; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014466:Ultraviolet Rays
chemical_list: D021343:Antifreeze Proteins, Type III; D003374:Coumarins; C092813:RD3 protein, eel; C053769:4-bromomethyl-6,7-dimethoxycoumarin