DEVELOPING SOFTWARE TOOLS FOR STRUCTURE DETERMINATION OF LARGE PROTEINS BY NMR SPECTROSCOPY

Abstract

Resonance assignment is often the most challenging step in NMR studies of protein structure, especially for large proteins. Although deuteration and TROSY techniques allow the assignment of backbone and 13CI? resonances for proteins up to 100 kDa, deuteration also severely reduces the number of NOE-derived distance constraints. For accurate structure determination, it is important to assign side-chain resonances in protonated large proteins.In this study, a software tool was developed to facilitate the assignment of aliphatic side-chain resonances in uniformly 13C,15N-labeled large proteins. It adopts a general strategy recently proposed by our lab, which makes use of 4D 13C,15N-edited NOESY, 3D MQ-(H)CCmHm-TOCSY, and prior backbone and 13CI? assignments. Not only can it greatly accelerate the assignment process, it also allows more resonances at distal end of side-chains to be assigned from weak NOEs, hence providing more distance constraints for accurate structure determination of large proteins.