The NMR determination of the structure of large biological macromolecules in solution is primarily limited by fast transverse relaxation that broadens lines and reduces spectral resolution. Several steps ahead have been made recently to overcome this limitation. Direct detection of heteronuclei, and of 13C in particular, offers a valuable alternative to 1H detection. However, 13C direct detection has not been widely applied to study biological macromolecules. The availability of increasingly high magnetic fields and the advent of cryogenically-cooled probeheads have nowadays moved the sensitivity of NMR spectroscopy into regions that were unforeseeable less than a decade ago. As a consequence, the 13C sensitivity has been dramatically increased up to a level suitable to turn 13C detected experiments into a routine tool for biomolecular NMR applications. We will discuss here instrumental aspects and we will propose various schemes to face the problem of the large homonuclear one-bond carbon-carbon couplings that evolve during the acquisition delay. We will then summarize the set of experiments based exclusively on heteronuclei that allows one to perform the complete sequence specific assignment of a 13C, 15N labelled protein without using 1H excitation and detection. As far as paramagnetism-based structural constraints are concerned, it will be shown that 13C NMR studies of paramagnetic molecules provide in principle the same content of information as 1H studies, but the loss of information due to paramagnetism-induced line broadening will be much less effective. Finally, an analytical list of pulse sequences for 13C protonless NMR spectroscopy will be given.

13C detected protonless NMR spectroscopy of proteins in solution / W.Bermel; I.Bertini; I.C.Felli; M.Piccioli; R.Pierattelli. - In: PROGRESS IN NUCLEAR MAGNETIC RESONANCE SPECTROSCOPY. - ISSN 0079-6565. - STAMPA. - 48:(2006), pp. 25-45. [10.1016/j.pnmrs.2005.09.002]

13C detected protonless NMR spectroscopy of proteins in solution

BERTINI, IVANO;FELLI, ISABELLA CATERINA;PICCIOLI, MARIO;PIERATTELLI, ROBERTA
2006

Abstract

The NMR determination of the structure of large biological macromolecules in solution is primarily limited by fast transverse relaxation that broadens lines and reduces spectral resolution. Several steps ahead have been made recently to overcome this limitation. Direct detection of heteronuclei, and of 13C in particular, offers a valuable alternative to 1H detection. However, 13C direct detection has not been widely applied to study biological macromolecules. The availability of increasingly high magnetic fields and the advent of cryogenically-cooled probeheads have nowadays moved the sensitivity of NMR spectroscopy into regions that were unforeseeable less than a decade ago. As a consequence, the 13C sensitivity has been dramatically increased up to a level suitable to turn 13C detected experiments into a routine tool for biomolecular NMR applications. We will discuss here instrumental aspects and we will propose various schemes to face the problem of the large homonuclear one-bond carbon-carbon couplings that evolve during the acquisition delay. We will then summarize the set of experiments based exclusively on heteronuclei that allows one to perform the complete sequence specific assignment of a 13C, 15N labelled protein without using 1H excitation and detection. As far as paramagnetism-based structural constraints are concerned, it will be shown that 13C NMR studies of paramagnetic molecules provide in principle the same content of information as 1H studies, but the loss of information due to paramagnetism-induced line broadening will be much less effective. Finally, an analytical list of pulse sequences for 13C protonless NMR spectroscopy will be given.
48
25
45
W.Bermel; I.Bertini; I.C.Felli; M.Piccioli; R.Pierattelli
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2158/309774
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