3-DIMENSIONAL N-15-H-1-H-1 AND N-15-C-13-H-1 NUCLEAR-MAGNETIC-RESONANCE STUDIES OF HPR, A CENTRAL COMPONENT OF THE PHOSPHOENOLPYRUVATE-DEPENDENT PHOSPHOTRANSFERASE SYSTEM FROM ESCHERICHIA-COLI - ASSIGNMENT OF BACKBONE RESONANCES

NAJ VANNULAND, AA VANDIJK, K DIJKSTRA, FHJ VANHOESEL, RM SCHEEK, GT ROBILLARD

OnderzoeksoutputAcademicpeer review

Samenvatting

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 N-15 and C-13 to overcome spectral overlap. Complete assignments were obtained for the backbone H-1, N-15 and C-13 resonances, using three-dimensional heteronuclear H-1 NOE (H-N)-H-1-N-15 multiple-quantum coherence spectroscopy (3D-NOESY-HMQC) and three-dimensional heteronuclear total correlation (H-N)-H-1-N-15 multiple-quantum coherence spectroscopy (3D-TOCSY-HMQC) experiments on N-15-enriched HPr and an additional three-dimensional triple-resonance (HN-N-C)-H-1-N-15-C-13-alpha correlation spectroscopy (HNCA) experiment on C-13, N-15-enriched HPr. Many of the sequential backbone H-1 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.

Originele taal-2English
Pagina's (van-tot)483-491
Aantal pagina's9
TijdschriftEuropean Journal of Biochemistry
Volume203
Nummer van het tijdschrift3
StatusPublished - 1-feb-1992

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