Folding Kinetics of Staphylococcal Nuclease Studied by Tryptophan Engineering and Rapid Mixing Methods
نویسندگان
چکیده
منابع مشابه
Early events during folding of wild-type staphylococcal nuclease and a single-tryptophan variant studied by ultrarapid mixing.
A continuous-flow mixing device with a dead time of 100 micros coupled with intrinsic tryptophan and 1-anilinonaphthalene-8-sulfonate (ANS) fluorescence was used to monitor structure formation during early stages of the folding of staphylococcal nuclease (SNase). A variant with a unique tryptophan fluorophore in the N-terminal beta-barrel domain (Trp76 SNase) was obtained by replacing the singl...
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Staplhylococcal nuclease undergoes a reversible structural transition between (p)h3 and 4 which be mesured by changes in tryptoham fluorescence. A stopped-flow spectrofluorometer was used to study the kinetics renaturation of nuclease from the acidified form on neutralization, the refolding is fast and the data can be described as a sequence of two first-order processes with half times of about...
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Information on the time-dependence of molecular species is critical for elucidating reaction mechanisms in chemistry and biology. Rapid flow experiments involving turbulent mixing of two or more solutions continue to be the main source of kinetic information on protein folding and other biochemical processes, such as ligand binding and enzymatic reactions. Recent advances in mixer design and de...
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FT-IR spectroscopic, small-angle X-ray scattering and calorimetric measurements have been applied to explore the effect of the macromolecular crowder agent Ficoll on the temperature- and pressure-dependent stability diagram and folding reaction of the protein Staphylococcal Nuclease (SNase). Additionally, we compare the experimental data with approximate theoretical predictions. We found that t...
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2.1. Turbulent Mixing 1838 2.2. Stopped-Flow Methods 1838 2.3. Continuous-Flow Techniques 1839 3. Detection Methods 1841 3.1. Tryptophan Fluorescence 1841 3.2. ANS Fluorescence 1842 3.3. Fluorescence Energy Transfer 1842 3.4. Continuous-Flow Absorbance 1843 3.5. Probes of Secondary Structure Formation 1844 3.6. Small-Angle X-ray Scattering 1844 3.7. Other Detection Methods 1844 4. Quenched-Flow...
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ژورنال
عنوان ژورنال: Journal of Molecular Biology
سال: 2007
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2007.02.006