期刊
JOURNAL OF PHYSICAL CHEMISTRY B
卷 117, 期 17, 页码 5009-5018出版社
AMER CHEMICAL SOC
DOI: 10.1021/jp402946c
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资金
- National Science Foundation (NSF) [0832584, CHE-9629688, CHE-9208463, DMB-8415048, OLA-9977486, BIR-9214394]
- National Institutes of Health (NIH) [T32 GM08293, 1 S10 R1U3866-01, P41RR02301, P41GM66326, RR02781, RR08438]
- NIDDK [DK79895]
- University of Wisconsin
- USDA
- NIGMS [P41-GM103311]
- Division Of Chemistry
- Direct For Mathematical & Physical Scien [0832584] Funding Source: National Science Foundation
A series of non-natural:infrared probes is reported that consist of a metal-tricarbonyl modified with a -(CH2)(n)- linker and cysteine-specific leaving group. They can be site specifically attached to proteins using mutagenesis and similar protocols for EPR spin labels, which have the same leaving group. We characterize the label's frequencies. and lifetimes. using 2D IR spectroscopy in solvents of varying dielectric The frequency range spans 10 cm(-1) and the variation in lifetimes ranges from 6 to 19 Ps, indicating that these probes are very sensitive to their environments. Also, We attached probes with -(CH2)- -(CH2)(3)-, and -(CH2)(4)- linkers to ubiquitin at positions 6 and 63 and collected spectra in aqueous huller. The frequencies and lifetimes were correlated for 3C and 4C linkers, as they were in the solvents, but did not correlate for the 1C linker We conclude that lifetime Measures solvation, whereas frequency reflects the electrostatics of the environment, which in the case of the 1C linker is a measure of the protein electrostatic field. We also labeled V71C alpha-synuclein in buffer and membrane-bound Unlike most other infrared labels, this label has extremely strong cross sections and thus can be measured with 2D IR spectroscopy at sub-millimolar concentrations. We expect that these labels will find use in studying the structure and dynamics of membrane bound, aggregated, and kinetically evolving proteins for which high signal-to-noise at low protein Concentrations is imperative.
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