RESUMO
We report the determination of the global fold of human ubiquitin using protein backbone NMR residual dipolar coupling and long-range nuclear Overhauser effect (NOE) data as conformational restraints. Specifically, by use of a maximum of three backbone residual dipolar couplings per residue (Ni-H N i, Ni-C'(i-1), H N i - C'(i-1)) in two tensor frames and only backbone H N -H N NOEs, a global fold of ubiquitin can be derived with a backbone root-mean-square deviation of 1.4 A with respect to the crystal structure. This degree of accuracy is more than adequate for use in databases of structural motifs, and suggests a general approach for the determination of protein global folds using conformational restraints derived only from backbone atoms.
Assuntos
Ressonância Magnética Nuclear Biomolecular/métodos , Ubiquitina/química , Sequência de Aminoácidos , Humanos , Magnetismo , Estrutura Terciária de Proteína , Proteínas/químicaRESUMO
The implementation of [13Calpha,13C',15N,2Halpha] labelled amino acids into proteins allows the acquisition of high resolution triple resonance experiments. We present for the first time resonance assignments facilitated by this new labelling strategy. The absence of 1JCalpha,Cbeta couplings enables us to measure 1JCalpha,C' scalar and 1DCalpha,C' residual dipolar coupling constants using modified HNCA experiments which do not suffer from sensitivity losses characteristic for 13C constant time experiments.
