64, JQ = 25 Hz). The LY2835219 solubility dmso C12E6 and n-hexanol were purchased from Sigma–Aldrich and used without further purification. The variants of the HSQC sequence were tested on a sample of d-sucrose (3) (30 mg) dissolved in 500 μl D2O. For all measurements the nominal temperature was set to 298 K unless indicated otherwise. All F2-coupled CLIP/CLAP-HSQC spectra were acquired with a high spectral resolution of 0.3 Hz/point, for accurate measurement of small residual dipolar couplings. The

15N–1H pure shift HSQC spectrum was recorded for 1.6 mM [U–15N]–Penicillium antifungal protein (PAF) (95%: 5% H2O:D2O), pH 5.0, at 300 K. Spectra were recorded with a proton 90° pulse of 15 μs, a carbon 90° pulse of 15.7 μs for acquisition, a carbon 90° pulse of 80.0 μs for GARP decoupling, smoothed chirp pulses (Crp60,0.5,20.1)

of 500 μs duration for broadband 13C inversion and (Crp60comp.4) of 2 ms for broadband 13C refocusing. 1H–15N HSQC spectra were collected with nitrogen 90° pulses of 29 μs for acquistion and 250 μs for WALTZ16 decoupling. For processing the 3D raw data sets acquired with the pulse sequences presented, a Bruker AU program (available at http://nmr.chemistry.manchester.ac.uk) was used to reconstruct the 2D interferograms. Prior to 2D Fourier transformation, the data were apodized by multiplying with a 90° shifted sine-squared function and then zero-filled by a factor of two in both dimensions, to yield a spectral resolution of 0.3–0.5 Hz/point in the 1H dimension. Due to the increasing interest in the use of RDCs in recent years, numerous Idelalisib methods based on measuring frequency differences between multiplet components have been developed for the measurement of one-bond heteronuclear coupling constants. The check details most widely used approach is based on the HSQC experiment, with heteronuclear couplings retained in the F1 or F2 dimension. To circumvent spectral crowding due to the increased number of cross-peaks in the coupled spectra, E-COSY [12], spin-state selective [13], [14] and [15], IPAP [16] and TROSY [17], [18] and [19] methods have been proposed. Unfortunately, all these methods

suffer from additional splittings of cross-peaks due to the co-evolution during data acquisition of coupling interactions other than the desired heteronuclear one-bond coupling. To eliminate line-splittings caused by multiple bond heteronuclear couplings in the F1-coupled HSQC sequence, a gradient enhanced BIRD(r) module has been employed during the evolution period t1, yielding simplified cross peaks with only splittings due to the desired one-bond couplings in the F1 dimension [20] and [21]. However, heteronuclear correlation experiments coupled in the indirect F1 dimension are limited by the necessity of acquiring large numbers of t1 points to achieve sufficiently high digital resolution, therefore making the experiment rather time-consuming.