2012
DOI: 10.1021/ja307606f
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Fast Photochemical Oxidation of Proteins and Mass Spectrometry Follow Submillisecond Protein Folding at the Amino-Acid Level

Abstract: We report a study of sub-millisecond protein folding with amino-acid residue resolution achieved with a two-laser pump/probe experiment with analysis by mass spectrometry. The folding of a test protein, barstar, can be triggered by a laserinduced temperature jump (T jump) from ~0 °C to ~ room temperature. Subsequent reactions via FPOP (fast photochemical oxidation of proteins) at various fractional millisecond points after the T jump leads to oxidative modification of solvent-accessible side chains whose “prot… Show more

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Cited by 98 publications
(126 citation statements)
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“…FPOP of acid-unfolded apo-Mb produces F U = 4.3%, much lower than the expected value of 25% (Figure 5b). Similarly, FPOP of semi-unfolded barstar with EVF = 25% [63] resulted in F U = 7% (Figure 2a in reference [41]). It is concluded that F U values in FPOP tend to be too low (i.e., the percentage of oxidized protein is higher than expected).…”
Section: Fpop Mass Spectramentioning
confidence: 88%
“…FPOP of acid-unfolded apo-Mb produces F U = 4.3%, much lower than the expected value of 25% (Figure 5b). Similarly, FPOP of semi-unfolded barstar with EVF = 25% [63] resulted in F U = 7% (Figure 2a in reference [41]). It is concluded that F U values in FPOP tend to be too low (i.e., the percentage of oxidized protein is higher than expected).…”
Section: Fpop Mass Spectramentioning
confidence: 88%
“…Hydrogen-deuterium exchange MS (HDX-MS) is particularly suited to measure secondary and tertiary structure stability through backbone exchange, whereas HRF-MS has been effective at measuring the relative solvent accessibility of specific amino acid side chains mediated by intramolecular tertiary and intermolecular quaternary structure interactions. Hydroxyl radicals can be generated by a variety of methods in each case the chemistry has been shown to be quite similar and the radicals react with side chains of surface residues resulting in well characterized oxidation products (7,10,11). As up to 18 side chains are potential probes, the overall protein coverage and resolution of the method is theoretically high.…”
Section: Hydroxyl Radical Footprinting (Hrf)mentioning
confidence: 99%
“…Three solution-phase labelling methods, i.e. hydroxyl radical footprinting [28][29][30], cross-linking [31][32][33] and HDX-MS [10,[34][35][36][37], have been extensively reviewed. Here, each technique is briefly described.…”
Section: Solution-phase Ms-based Techniquesmentioning
confidence: 99%