Structure Solution of Nano-Crystalline Small Molecules Using MicroED and Solid-State NMR Dipolar-Based Experiments

Three-dimensional electron diffraction crystallography (microED) can solve structures of sub-micrometer crystals, which are too small for single crystal X-ray crystallography. However, R factors for the microED-based structures are generally high because of dynamic scattering. That means R factor may not be reliable provided that kinetic analysis is used. Consequently, there remains ambiguity to locate hydrogens and to assign nuclei with close atomic numbers, like carbon, nitrogen, and oxygen. Herein, we employed microED and ssNMR dipolar-based experiments together with spin dynamics numerical simulations. The NMR dipolar-based experiments were H-1-N-14 phase-modulated rotational-echo saturation-pulse double-resonance (PM-S-RESPDOR) and H-1-H-1 selective recoupling of proton (SERP) experiments. The former examined the dephasing effect of a specific H-1 resonance under multiple H-1-N-14 dipolar couplings. The latter examined the selective polarization transfer between a H-1-H-1 pair. The structure was solved by microED and then validated by evaluating the agreement between experimental and calculated dipolar-based NMR results. As the measurements were performed on H-1 and N-14, the method can be employed for natural abundance samples. Furthermore, the whole validation procedure was conducted at 293 K unlike widely used chemical shift calculation at 0 K using the GIPAW method. This combined method was demonstrated on monoclinic l-histidine.

Automated summary

MicroED is used to solve structures of sub-micrometer crystals, but the high R factor makes it difficult to locate hydrogens and assign nuclei with close atomic numbers. By combining microED and ssNMR experiments with spin dynamics numerical simulations, this study provides a method for natural abundance samples.