Broadband Heteronuclear Solid-State NMR Experiments by Exponentially Modulated Dipolar Recoupling without Decoupling

Abstract

We present a novel solid-state NMR method for heteronuclear dipolar recoupling without decoupling. The method, which introduces the concept of exponentially modulated rf fields, provides efficient broadband recoupling with large flexibility with respect to hetero- or homonuclear applications, sample spinning frequency, and operation without the need for high-power (1)H decoupling. For previous methods, the latter has been a severe source of sample heating which may cause detoriation of costly samples. The so-called EXPonentially mOdulated Recoupling Technique (EXPORT) is described analytically and numerically, and demonstrated experimentally by 1D (13)C spectra and 2D (13)C-(15)N correlation spectra of (13)C,(15)N-labeled samples of GB1, ubiquitin, and fibrils of the SNNFGAILSS fragment of amylin. Through its flexible operation, robustness, and strong performance, it is anticipated that EXPORT will find immediate application for both hetero- and homonuclear dipolar recoupling in solid-state NMR of (13)C,(15)N-labeled proteins and compounds of relevance in chemistry.

Figure 1

(a) The EXPORT pulse sequence for heteronuclear dipolar recoupling without decoupling embedded in a typical 2D NCO/NCA chemical shift correlation experiment. (b) The amplitude (red) and phase (blue) modulation schemes of the basic elements with the ¹³C and ¹⁵N fields represented by solid and broken lines, respectively, for EXPORT with C=6ω_r, B_I=3ω_r/8, B_S=5ω_r/8, and ω_r/2π=12 kHz.

Figure 2

¹⁵N to ¹³C_α coherence transfer efficiencies calculated for EXPORT using the parameters in Fig. 1b. (a–c) 2D ¹⁵N vs ¹³C offset plots for EXPORT (c), an ^OCNCA optimal control sequence (b), and DCP (a). (d–f) 2D ¹⁵N vs ¹³C rf field strength plots (scaling factors relative to the nominal values) for DCP (d), EXPORT with high digitization of the rf field (100 points over 1 rotor period) (e), and EXPORT with lower digitization (20 points over 1 rotor period) (f). Simulations were made using SIMPSON with parameters for a directly bonded ¹⁵N-¹³C_α spin system, powder averaging with 5 γ_CR and 144 REPULSION angles, and a spinning frequency of 12 kHz at 16.4 T. Broken lines mark carrier frequencies.

Figure 3

Experimental (signal integrals for the target spin spectral region) and simulated ¹⁵N to ¹³C’ (a) and ¹⁵N to ¹³C_α (b) coherence transfer efficiencies for DCP (experiment: red crosses, simulation: red line) and EXPORT (experiment: blue circles, simulation: blue line) as function of the ¹³C rf field strength mismatch. Experimental spectra were obtained for U-¹³C,¹⁵N-labeled FGAIL in a SNNFGAILSS fibril sample. Representative ¹³C spectra following (c) ¹⁵N to ¹³C_α and (d) ¹⁵N to ¹³C’ transfer for EXPORT (left) and DCP (right). In accord with Fig. 1a, the spectra were recorded using CP for the initial ¹H-¹⁵N transfer and SPINAL-64 decoupling³² (80 kHz) was used during acqusition. All spectra were recorded at 11.9 kHz spinning with carrier frequencies at 120 ppm for ¹⁵N and 50/172 ppm for NCA/NCO transfer. DCP used ω_rf,C/2π = 50.2 kHz, ω_rf,N/2π = 39.3 kHz, and 120 kHz ¹H decoupling. EXPORT used C=7ω_r, B_I=3ω_r/8, B_S=5ω_r/8, and no ¹H decoupling.

Figure 4

Experimental spectra for U-¹³C,¹⁵N-labeled samples of (a) GB1 and (b) ubiquitin recorded at 16.4 T using EXPORT for ¹⁵N-¹³C transfer with C =3ω_r, B_I=3ω_r/8, B_S=5ω_r/8, and τ_mix=2.35 ms. In accord with Fig. 1a, the spectra were recorded using CP for the initial ¹H-¹⁵N transfer. SPINAL-64 decoupling (70 kHz) was used in the detection periods. The 2D spectrum (a) used 23.81 kHz sample spinning without ¹H decoupling during EXPORT, 560 points in the indirect and 4096 point in the direct dimensions. The 1D spectra (b) used 10.02 kHz spinning, carrier frequencies at 40 (top), 120 (middle), and 180 (bottom) ppm, and 90 kHz ¹H decoupling during EXPORT.