Abstract
Virtually complete sequence specific 1H and 15N resonance assignments are presented for acid denatured reduced E. coli glutaredoxin 3. The sequential resonance assignments of the backbone rely on the combined use of 3D F1-decoupled ROESY-15N-HSQC and 3D 15N-HSQC-(TOCSY-NOESY)-15N-HSQC using a single uniformly 15N labelled protein sample. The sidechain resonances were assigned from a 3D TOCSY-15N-HSQC and a homonouclear TOCSY spectrum. The presented assignment strategy works in the absence of chemical exchange peaks with signals from the native conformation and without 13C/15N double labelling. Chemical shifts, 3J(αH, NH) coupling constants and NOEs indicate extensive conformational averaging of both backbone and side chains in agreement with a random coil conformation. The only secondary structure element persisting at pH 3.5 appears to be a short helical segment comprising residues 37 to 40.
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Abbreviations
- HSQC:
-
heteronuclear single quantum coherence
- NMR:
-
nuclear magnetic resonance
- NOE:
-
nuclear Overhauser effect
- NOESY:
-
two-dimensional NOE spectroscopy
- ROE:
-
nuclear Overhauser effect in the rotating frame
- ROESY:
-
two-dimensional ROE spectroscopy
- TOCSY:
-
total correlation spectroscopy
- TPPI:
-
time proportional phase incrementation
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Correspondence to: G. Otting
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Nordstrand, K., Ponstingl, H., Holmgren, A. et al. Resonance assignment and structural analysis of acid denatured E. coli [U-15N]-glutaredoxin 3. Eur Biophys J 24, 179–184 (1996). https://doi.org/10.1007/BF00180275
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DOI: https://doi.org/10.1007/BF00180275