%0 Journal Article
%A Norton-Baker, Brenna
%A Mehrabi, Pedram
%A Boger, Juliane
%A Schoenherr, Robert Frank
%A von Stetten, David
%A Schikora, Hendrik
%A Kwok, Ashley O.
%A Martin, Rachel W.
%A Miller, R. J. Dwayne
%A Redecke, Lars
%A Schulz, Eike C.
%T A simple vapor-diffusion method enables protein crystallization inside the HARE serial crystallography chip
%J Acta crystallographica / Section D
%V 77
%N 6
%@ 2059-7983
%C Bognor Regis
%I Wiley
%M PUBDB-2021-02689
%P 820 - 834
%D 2021
%X Fixed-target serial crystallography has become an important method for the study of protein structure and dynamics at synchrotrons and X-ray free-electron lasers. However, sample homogeneity, consumption and the physical stress on samples remain major challenges for these high-throughput experiments, which depend on high-quality protein microcrystals. The batch crystallization procedures that are typically applied require time- and sample-intensive screening and optimization. Here, a simple protein crystallization method inside the features of the HARE serial crystallography chips is reported that circumvents batch crystallization and allows the direct transfer of canonical vapor-diffusion conditions to in-chip crystallization. Based on conventional hanging-drop vapor-diffusion experiments, the crystallization solution is distributed into the wells of the HARE chip and equilibrated against a reservoir with mother liquor. Using this simple method, high-quality microcrystals were generated with sufficient density for the structure determination of four different proteins. A new protein variant was crystallized using the protein concentrations encountered during canonical crystallization experiments, enabling structure determination from ∼55 µg of protein. Additionally, structure determination from intracellular crystals grown in insect cells cultured directly in the features of the HARE chips is demonstrated. In cellulo crystallization represents a comparatively un­explored space in crystallization, especially for proteins that are resistant to crystallization using conventional techniques, and eliminates any need for laborious protein purification. This in-chip technique avoids harvesting the sensitive crystals or any further physical handling of the crystal-containing cells. These proof-of-principle experiments indicate the potential of this method to become a simple alternative to batch crystallization approaches and also as a convenient extension to canonical crystallization screens.
%F PUB:(DE-HGF)16
%9 Journal Article
%$ pmid:34076595
%U <Go to ISI:>//WOS:000659143800009
%R 10.1107/S2059798321003855
%U https://bib-pubdb1.desy.de/record/459716