EPR‐Based Approach for the Localization of Paramagnetic Metal Ions in Biomolecules

Abstract

Metal ions play an important role in the catalysis and folding of proteins and oligonucleotides. Their localization within the three-dimensional fold of such biomolecules is therefore an important goal in understanding structure-function relationships. A trilateration approach for the localization of metal ions by means of long-range distance measurements based on electron paramagnetic resonance (EPR) is introduced. The approach is tested on the Cu(2+) center of azurin, and factors affecting the precision of the method are discussed.

Distance measurements between manganese(ii) and nitroxide spin-labels by DEER determine a binding site of Mn2+ in the HP92 loop of ribosomal RNA

“…It has also been successfully applied to determine the distances between other types of paramagnetic centers in biomolecules, 8 such as Cu 2+ -Cu 2+ , 9,10 Gd 3+ -Gd 3+ , [11][12][13] Mn 2+ -Mn 2+ , 14 pairs of iron-sulfur clusters 15,16 and trityl-trityl radicals. 17,18 Biomolecular heterospin label distance measurements, including nitroxideÀCu 2+ , [19][20][21] nitroxide-iron-sulfur cluster 22 and nitroxide-Gd 3+ , 23,24 have also been reported. A recent important alternative application of DEER, in combination with site directed spin labelling, has been locating metal ion binding sites by means of paramagnetic metal ionnitroxide distance measurements in a biomolecule, as has been demonstrated on Cu 2+ binding sites.…”

“…17,18 Biomolecular heterospin label distance measurements, including nitroxideÀCu 2+ , [19][20][21] nitroxide-iron-sulfur cluster 22 and nitroxide-Gd 3+ , 23,24 have also been reported. 19,21 Metal ions play a crucial role in RNA structure and function. 19,21 Metal ions play a crucial role in RNA structure and function.…”

“…19,21 Metal ions play a crucial role in RNA structure and function. It was found to be preferably situated in the minor groove of the double strand region close to the HP92 loop.…”

“…19,21 Metal ions play a crucial role in RNA structure and function. 4-7 The most common application of DEER is to measure distances between two nitroxide spin labels attached at specific points of interest in a biomolecule.…”

The contribution of modern EPR to structural biology

“…If the ligand can be spin-labelled or the metal ion is paramagnetic, they can be localized by a distance matrix approach [36] or by an approach akin to the global positioning system (GPS), where the position of an observer can be determined by at least three distance measurements to reference sites [37]. The distance matrix approach tests and relaxes the assumption of a rigid reference domain at the expense of additional measurements between reference points.…”