Three-dimensional 15N-1H-1H and 15N-13C-1H nuclear-magnetic resonance studies of HPr a central component of the phosphoenolpyruvate-dependent phosphotransferase system from Escherichia coli. Assignment of backbone resonances.

TitleThree-dimensional 15N-1H-1H and 15N-13C-1H nuclear-magnetic resonance studies of HPr a central component of the phosphoenolpyruvate-dependent phosphotransferase system from Escherichia coli. Assignment of backbone resonances.
Publication TypeJournal Article
Year of Publication1992
Authorsvan Nuland, N. A. J., A. A. van Dijk, K. Dijkstra, F. H. van Hoesel, R. M. Scheek, and G. T. Robillard
JournalEur J Biochem
Volume203
Issue3
Pagination483-91
Date Published1992 Feb 1
ISSN0014-2956
KeywordsBacterial Proteins, Carbon Isotopes, Escherichia coli, Magnetic Resonance Spectroscopy, Nitrogen Isotopes, Phosphoenolpyruvate Sugar Phosphotransferase System, Protein Conformation
Abstract

We have performed three-dimensional NMR studies on a central component of the phosphoenolpyruvate-dependent phosphotransferase system of Escherichia coli, denoted as HPr. The protein was uniformly enriched with 15N and 13C to overcome spectral overlap. Complete assignments were obtained for the backbone 1H, 15N and 13C resonances, using three-dimensional heteronuclear 1H NOE 1H-15N multiple-quantum coherence spectroscopy (3D-NOESY-HMQC) and three-dimensional heteronuclear total correlation 1H-15N multiple-quantum coherence spectroscopy (3D-TOCSY-HMQC) experiments on 15N-enriched HPr and an additional three-dimensional triple-resonance 1HN-15N-13C alpha correlation spectroscopy (HNCA) experiment on 13C, 15N-enriched HPr. Many of the sequential backbone 1H assignments, as derived from two-dimensional NMR studies [Klevit, R.E., Drobny, G.P. & Waygood, E.B. (1986) Biochemistry 25, 7760-7769], were corrected. Almost all discrepancies are in the helical regions, leaving the published antiparallel beta-sheet topology almost completely intact.

Alternate JournalEur. J. Biochem.
PubMed ID1735433