Quantitative Analysis of Solid-State Homonuclear Correlation Spectra of Antiparallel β‑Sheet Alanine Tetramers

Poly-l-alanine (PLA) sequences are a key element in the structure of the crystalline domains of spider dragline silks, wild silkworm silks, antifreeze proteins, and amyloids. To date, no atomic-level structures of antiparallel (AP)-PLA longer than Ala<sub>4</sub> have been reported using the single-crystal X-ray diffraction analysis. In this work, dipolar-assisted rotational resonance solid-state NMR spectra were observed to determine the effective internuclear distances of <sup>13</sup>C uniformly labeled alanine tetramer with antiparallel (AP) β-sheet structure whose atomic coordinates are determined from the X-ray crystallographic analysis. Initial build-up rates, <i>R</i><sub><i>j</i>,<i>k</i></sub>, were obtained from the build-up curves of the cross peaks by considering the internuclear distances arising in the master equation. Subsequently, experimentally obtained effective internuclear distances, <i>r</i><sup>eff</sup><sub><i>j</i>,<i>k</i></sub>(obs), were compared with the calculated <i>r</i><sup>eff</sup><sub><i>j</i>,<i>k</i></sub>(calc) values obtained from the X-ray crystallographic data. Fairly good correlation between <i>r</i><sup>eff</sup><sub><i>j</i>,<i>k</i></sub>(obs) and <i>r</i><sup>eff</sup><sub><i>j</i>,<i>k</i></sub>(calc) was obtained in the range of 1.0–6.0 Å, with the standard deviation of 0.244 Å, without considering the zero-quantum line-shape functions. It was further noted that the internuclear distances of intermolecular contributions provide details relating to the molecular packing in solid-state samples. Thus, the present data agree well with AP-β-sheet packing but do not agree with P-β-sheet packing.