Generating Accurate Contact Maps of Transient Long-Range Interactions in Intrinsically Disordered Proteins by Paramagnetic Relaxation Enhancement
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland. Electronic address: .Biophysical Journal (Impact Factor: 3.97). 04/2013; 104(8):1635-6. DOI: 10.1016/j.bpj.2013.01.060
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ABSTRACT: The single His-15 of hen egg lysozyme reacts with 2,2,6,6-tetramethyl-4-(bromoacetamido)piperidinyl-1-oxy or 2,2,5,5-tetramethyl-3-(bromoacetamido)pyrrolidinyl-1-oxy to give a spin-labeled enzyme [Wien, R. W., Morrisett, J. D., & McConnell, H. M. (1972) Biochemistry 11, 3707-3716]. High-field 1H NMR spectra (300 and 500 MHz) of these species in 2H2O contain protein peaks selectively broadened by dipolar coupling to the unpaired electron spin. While usually difficult to discern in the spectrum itself, broadened resonances are revealed in difference spectra obtained by subtracting the original spectrum from one taken after reduction of the nitroxide radical with ascorbate. The heights of difference spectra peaks are related in a simple way to r-6, where r is the label to proton distance. These distances were used to solve for the location of the electron spin by using algorithms from distance geometry. The spin was found to lie in a hydrophobic groove between Phe-3 and Asp-87. These results demonstrate the feasibility of spin-labeling for accurate distance measurements in proteins through the use of distance geometry.Biochemistry 08/1984; 23(18):4261-6. DOI:10.1021/bi00313a038 · 3.01 Impact Factor
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ABSTRACT: In the last decade it has become evident that disordered states of proteins play important physiological and pathological roles and that the transient tertiary interactions often present in these systems can play a role in their biological activity. The structural characterization of such states has so far largely relied on ensemble representations, which in principle account for both their local and global structural features. However, these approaches are inherently of low resolution due to the large number of degrees of freedom of conformational ensembles and to the sparse nature of the experimental data used to determine them. Here, we overcome these limitations by showing that tertiary interactions in disordered states can be mapped at high resolution by fitting paramagnetic relaxation enhancement data to a small number of conformations, which can be as low as one. This result opens up the possibility of determining the topology of cooperatively collapsed and hidden folded states when these are present in the vast conformational landscape accessible to disordered states of proteins. As a first application, we study the long-range tertiary interactions of acid-unfolded apomyoglobin from experimentally measured paramagnetic relaxation enhancement data.Biophysical Journal 04/2013; 104(8):1740-51. DOI:10.1016/j.bpj.2013.02.019 · 3.97 Impact Factor
Article:  Spin labeling of proteinsMethods in Enzymology 02/1989; 177:86-121. DOI:10.1016/0076-6879(89)77007-5 · 2.19 Impact Factor
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