Rigaku Symposium at Yale University

Microcrystal electron diffraction (MicroED/3D ED), which solves atomic structures from crystals in the sub-micron size range using electron radiation, has evolved from an exploratory technique into a cutting-edge analytical method practiced in laboratories worldwide. However, viewing MicroED solely as an extension of single-crystal diffraction, traditionally performed with X-rays on smaller crystals, underestimates its capabilities. With thousands of crystallites available simultaneously on a sample grid and measurements completed within minutes, we can obtain a more comprehensive view of our specimen by collecting crystallographic data from dozens or even hundreds of crystals. This approach allows for the analysis of powders containing mixtures of compounds, different polymorphs, or even small traces of contaminants, including ab-initio solutions, even in cases too complex for powder diffraction or where only minute sample amounts are available. In this contribution, we will discuss how high-throughput workflows in the Rigaku XtaLAB Synergy-ED electron diffractometer enable various beyond-single-crystal investigations with a high degree of automation and ease of use.

The Structural Science Facility (SSF) within Yale University's Chemical and Biophysical Instrumentation Center (CBIC) has recently acquired the Synergy-ED electron diffractometer. This instrument has already demonstrated its value across multiple applications, particularly in establishing stereochemistry directly from electron diffraction data. In this presentation, I will highlight several initial results obtained using the XtaLAB Synergy-ED, focusing specifically on workflows, best practices, and considerations for accurately determining absolute configurations.

Microcrystal Electron Diffraction (MicroED) has revolutionized structural biology by enabling atomic-resolution structure determination from nanocrystals too small for traditional X-ray methods. While often used for small molecules and peptides, the application of MicroED to macromolecules—proteins, membrane proteins, and complex biomolecules—has opened new frontiers.

In this talk, I’ll discuss how recent advancements in sample preparation, data collection, and processing have allowed us to push MicroED beyond peptides and small molecules into the realm of challenging macromolecular systems. Using case studies from recent work, we’ll explore what makes macromolecular MicroED different, why it works when it does, and what challenges remain. With new tools and techniques, could MicroED become a mainstream tool for protein structure determination? Let’s find out!

We introduce a newly developed micro-area X-ray diffractometer designed to deliver high-precision analysis for a wide range of sample types, including solid metals, ceramics, polymers, and thin films. The system features a 20 µm diameter X-ray beam generated by a high-brilliance micro-focus X-ray source combined with confocal multilayer mirrors, enabling localized diffraction measurements with exceptional accuracy. An open cradle with small sphere of confusion ensures precise sample positioning, while a large photon-counting hybrid pixel array detector covers a wide 2θ range for rapid data acquisition. Additionally, a high-resolution sample observation camera facilitates accurate beam alignment. The system supports various sample holders, accommodating fibers, films, bulk materials, and thin films, while the non-ambient and operando attachments enable in-situ XRD measurements with significantly faster acquisition times compared to conventional systems. This versatile and high-performance system redefines the capabilities of micro-area XRD, offering unprecedented flexibility and speed for cutting-edge materials research.

The material properties of polymers are profoundly influenced by their molecular microstructures, including monomer sequence, architecture, and tacticity. This lecture will present recent advances in precision polymer synthesis aimed at tailoring these microstructural features to achieve targeted properties in homopolymers and copolymers. Emphasis will be placed on the integration of advanced scattering techniques to elucidate structure–property relationships.  The presentation will begin with recent work from the speaker’s research group on the controlled synthesis and structural characterization of multicomponent bottlebrush block copolymers featuring sequence-defined backbones. Hierarchically assembled nanostructures were confirmed via small- and wide-angle X-ray scattering, with tunable characteristic length scales. The concept of precision synthesis will then be extended to the design of polymeric inhibitors for silica scale mitigation. A post-polymerization modification strategy enables high-throughput screening of pendant functionalities, facilitating the discovery of optimal structures for inhibition efficiency. Small-angle neutron scattering was employed to probe polymer chain conformation in aqueous solution—an essential parameter governing antiscaling performance.  Finally, the lecture will introduce a bimetallic molecular catalyst for stereocontrolled living radical polymerization. The influence of backbone stereoregularity on the resulting polymers’ thermomechanical and thermo-responsive properties will be discussed, as evaluated through a combination of X-ray scattering and various spectroscopic techniques.

By assembling materials such as metal-organic frameworks (MOFs) from molecular building blocks featuring reversible bonds, we can form new nanoscale and ultrathin materials whose properties are subject to exquisite spatiotemporal control. Anthracene photocycloaddition to dianthracene constitutes a particularly robust molecular switch that can be reliably reversed in the solid state. Our group prepares dianthracene ligands that, once incorporated into MOFs, can undergo thermal or light-driven retrocyclization to controllably alter properties such as dimensionality, fluorescence, porosity, and so on. This photo-driven retrocyclization can be used to precisely alter the in-plane properties of 2D materials, including at the ultrathin (few-nanometer) limit, allowing unprecedented control over the optical properties and solubility of exfoliated samples uniquely enabled by molecular materials design. 

Crystal structures are critical to unambiguously determine a compound’s identity, absolute configuration, molecular connectivity and crystal form, as well as whether polymorphs coexist or how the solid-state structure may explain a compound’s behavior and physical properties. X-ray crystallography is the most commonly used method to obtain crystal structures, but it can be limiting if samples with suitable crystallinity, dimensions and quantity are lacking. 

Electron diffraction offers a new solution by providing high-resolution structural insights from submicron-size samples. Scratching the bottom of a beaker to gather nanograms of a crystalline powder may suffice for electron diffraction study. In other words, electron diffraction allows for single-crystal crystallography to be performed on powder samples. This allows for solving crystal structures that were previously unsolvable, but also screening crystallization conditions even when trials yield only a small amount of precipitate and automating polymorphs screening over hundreds of nanocrystals spread on a TEM grid.

In this presentation, we will review the fundamentals of electron diffraction, the best practices for sample preparation and demonstrate how it is reshaping research in fields such as pharmaceuticals, metal-organic frameworks, synthetic chemistry and materials science. 

Understanding the solid form landscape of pharmaceuticals is crucial to the success of product development. Crystallization and crystal structure determination play a central role in this endeavor. While single-crystal X-ray diffraction is a go-to technique for insight to the solid state, it depends on the availability of sufficiently large crystals for the experiment. The introduction of micro electron diffraction to this effort offers expedient access to structural information and decreases the crystal size parameter for structure determination. This talk will share how the interplay of micro electron diffraction, single-crystal X-ray diffraction, and other techniques provides elucidation of the form space of new drug substances.

Structure determination of novel complex molecules by X-ray crystallography can be limited by crystal size. Electron diffraction (MicroED/3D ED) effectively overcomes this limitation, but can require careful interpretation of subtle features in the data to maximize the yield of structural information. This is especially true in light of the damaging effect of electrons on molecular crystals. New approaches in electron diffraction and new technologies, for data collection, detection and processing, allow for structures to be determined from arbitrary regions of nanocrystals and allows for the effects of electron-beam-induced radiation damage to be mapped and/or mitigated in nanocrystals. This in turn enables the application of these approaches for the discovery of new chemical matter and drug development.

MicroED stands to have a transformative impact on small molecule structure determination in the fields of natural products chemistry and complex molecule synthesis.  In this lecture I will present several recent projects from our group where MicroED has played an indispensable role in resolving the structures of complex natural products and advanced synthetic intermediates.  I will describe structural studies of lomaiviticin A, a highly oxidized cytotoxic bacterial metabolite.  The structure of lomaiviticin A was assigned by HRMS and NMR analysis, but remained essentially untested in the literature owing to the absence of a fully synthetic route to the isolate.  In 2019, we made the startling discovery, using MicroED, that the structure of lomaiviticin was incorrectly assigned.  Follow up studies that reveal the basis for the original misassignment will be described.  I will also highlight several synthetic intermediates from our group whose structure was uniquely solved by MicroED analysis.