Figure 3.

The implementation of the proposed strategy on $\mathit{\alpha }$ -synuclein renders NMR spectra so informative that proline resonances can be assigned just by visual inspection of the figure. To this end, the first ${}^{\mathrm{13}}\mathrm{C}{-}^{\mathrm{13}}\mathrm{C}$ planes of the 3D $\left(\mathrm{H}\right){\mathrm{CBCACON}}^{\mathrm{Pro}}$  (a), 3D $\left(\mathrm{H}\right){\mathrm{CCCON}}^{\mathrm{Pro}}$  (b), and 3D $\left(\mathrm{H}\right){\mathrm{CBCANCO}}^{\mathrm{Pro}}$  (c) are shown, and the ${}^{\mathrm{13}}\mathrm{C}{-}^{\mathrm{15}}\mathrm{N}$ plane of the 3D $\left(\mathrm{H}\right){\mathrm{CBCACON}}^{\mathrm{Pro}}$ is also shown (d). The portion of the primary sequence of $\mathit{\alpha }$ -synuclein hosting its five proline residues is also reported (below the spectra). The pulse sequence for acquiring the 3D $\left(\mathrm{H}\right){\mathrm{CBCACON}}^{\mathrm{Pro}}$ experiment (e) is reported as an example of the implementation of the proposed approach (dotted box). The delays are $\mathit{\epsilon }=t_{\mathrm{2}}\left(\mathrm{0}\right)$ , $\mathit{\delta }=\mathrm{3.6}$  ms, $\mathrm{\Delta }=\mathrm{9}$  ms, ${\mathrm{\Delta }}_{\mathrm{1}}=\mathrm{25}$  ms, ${\mathrm{\Delta }}_{\mathrm{2}}=\mathrm{8}$  ms, ${\mathrm{\Delta }}_{\mathrm{3}}={\mathrm{\Delta }}_{\mathrm{2}}$ – ${\mathrm{\Delta }}_{\mathrm{4}}=\mathrm{5.8}$  ms, and ${\mathrm{\Delta }}_{\mathrm{4}}=\mathrm{2.2}$  ms. The phase cycle is as follows: ${\mathit{\varphi }}_{\mathrm{1}}=$  x, -x; ${\mathit{\varphi }}_{\mathrm{2}}=$  8(x), 8(-x); ${\mathit{\varphi }}_{\mathrm{3}}=$  4(x), 4(-x); ${\mathit{\varphi }}_{\mathrm{4}}=$  2(x), 2(-x); ${\mathit{\varphi }}_{\mathrm{IPAP}}\left(\text{IP}\right)=$  x; and ${\mathit{\varphi }}_{\mathrm{IPAP}}\left(\text{AP}\right)=$  -y; ${\mathit{\varphi }}_{\mathrm{rec}}=$  x, -x, -x, x, -x, x, x, -x. Quadrature detection was obtained by incrementing phase ${\mathit{\varphi }}_{\mathrm{3}}\left(t_{\mathrm{1}}\right)$ and ${\mathit{\varphi }}_{\mathrm{4}}\left(t_{\mathrm{2}}\right)$ in a States–TPPI (time-proportional phase incrementation) manner. The IPAP (in-phase/antiphase) approach was implemented for homonuclear decoupling in the direct acquisition dimension to suppress the large one bond scalar coupling constants ( ${}^{\mathrm{1}}{J}_{\mathrm{C}\mathit{\alpha }-{\mathrm{C}}^{\prime }}$ ; Felli and Pierattelli, 2015); alternative approaches can be implemented that exploit band-selective homonuclear decoupling (Alik et al., 2020; Ying et al., 2014) or processing algorithms that, thus, only require the in-phase spectra (Karunanithy and Hansen, 2021; Shimba et al., 2003).