Comparative studies on manual and automatic backbone chemical shift assignments of 2H-13C-15N-labeled Ube2g1Reportar como inadecuado

Comparative studies on manual and automatic backbone chemical shift assignments of 2H-13C-15N-labeled Ube2g1 - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

Journal of Analytical Science and Technology

, 6:29

First Online: 16 October 2015Received: 06 July 2015Accepted: 27 September 2015


BackgroundUbe2g1, one of human E2 enzymes, possesses an additional acidic loop in the vicinity of the active Cys90 residue. The presence of the backbone chemical shifts bCSs is essential for various NMR studies of Ube2g1. Triple resonance experiments for the bCSs assignment of C-N-labeled Ube2g1 were not efficient due to the relatively high molecular size more than 20 kDa.

MethodsH-C-N-labeled Ube2g1 was prepared to increase the T2 relaxation time, and then the TROSY-based triple resonance spectra were recorded with H decoupling. After the bCSs of Ube2g1 were assigned manually, the resulting bCSs were compared with those from two different automatic assignment programs. The AutoAssign program that utilizes only the peak lists of the NMR spectra accomplished 69 % assignment of the bCSs. On the other hand, the RASPnmr program requires the additional reference CSs that can be predicted from a relevant PDB coordinate. The homologous models of Ube2g1 were generated by various modeling programs, and then their CSs were predicted by using the SHIFTX2 program.

ResultsThe implementations of the predicted bCSs into the RASPnmr analysis resulted in complete and accurate assignment of the bCSs of H-C-N-labeled Ube2g1.

ConclusionsThe reference bCSs calculated from various homologous models of Ube2g1 enabled the automatic bCSs assignment process by the RASPnmr program, and a similar application will be possible for the bCS assignments of other proteins.

KeywordsChemical shift assignment Deuteration Homologous modeling NMR RASPnmr SHIFTX2 Ube2g1 Yun-Seok Choi and Eun-Hee Kim contributed equally to this work.

Download fulltext PDF

Autor: Yun-Seok Choi - Eun-Hee Kim - Kyoung-Seok Ryu


Documentos relacionados