Impact of spin label rigidity on extent and accuracy of distance information from PRE data
 

Impact of spin label rigidity on extent and accuracy of distance information from PRE data

Abstract

Paramagnetic relaxation enhancement (PRE) is a versatile tool for NMR spectroscopic structural and kinetic studies in biological macromolecules. Here, we compare the quality of PRE data derived from two spin labels with markedly different dynamic properties for large RNAs using the I-A riboswitch aptamer domain (78 nt) from Mesoplamsa florum as model system. We designed two I-A aptamer constructs that were spin-labeled by noncovalent hybridization of short spin-labeled oligomer fragments. As an example of a flexible spin label, UreidoU-TEMPO was incorporated into the 3â?² terminal end of helix P1 while, the recently developed rigid spin-label Ã?m was incorporated in the 5â?² terminal end of helix P1. We determined PRE rates obtained from aromatic 13C bound proton intensities and compared these rates to PREs derived from imino proton intensities in this sizeable RNA (~78 nt). PRE restraints derived from both imino and aromatic protons yielded similar data quality, and hence can both be reliably used for PRE determination. For NMR, the data quality derived from the rigid spin label Ã?m is slightly better than the data quality for the flexible UreidoTEMPO as judged by comparison of the structural agreement with the I-A aptamer crystal structure (3SKI).



Source: Journal of Biomolecular NMR